PRIMER_TM_FORMULA=0
PRIMER_SALT_CORRECTIONS=0
PRIMER_SALT_DIVALENT=0.0
PRIMER_DNTP_CONC=0.0
PRIMER_LIB_AMBIGUITY_CODES_CONSENSUS=0
SEQUENCE_ID=thermod_default
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_THERMODYNAMIC_PARAMETERS_PATH=primer3_config/
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_ANY_TH=47.0
PRIMER_MAX_SELF_END_TH=47.0
PRIMER_MAX_HAIRPIN_TH=47.0
PRIMER_PAIR_MAX_COMPL_ANY_TH=47.0
PRIMER_PAIR_MAX_COMPL_END_TH=47.0
PRIMER_INTERNAL_MAX_SELF_ANY_TH=47.0
PRIMER_INTERNAL_MAX_SELF_END_TH=47.0
PRIMER_INTERNAL_MAX_HAIRPIN_TH=47.0
PRIMER_MAX_TEMPLATE_MISPRIMING_TH=47.0
PRIMER_PAIR_MAX_TEMPLATE_MISPRIMING_TH=47.0
PRIMER_WT_SELF_ANY_TH=0.0
PRIMER_WT_SELF_END_TH=0.0
PRIMER_WT_HAIRPIN_TH=0.0
PRIMER_WT_TEMPLATE_MISPRIMING_TH=0.0
PRIMER_INTERNAL_WT_SELF_ANY_TH=0.0
PRIMER_INTERNAL_WT_SELF_END_TH=0.0
PRIMER_INTERNAL_WT_HAIRPIN_TH=0.0
PRIMER_PAIR_WT_COMPL_ANY_TH=0.0
PRIMER_PAIR_WT_COMPL_END_TH=0.0
PRIMER_PAIR_WT_TEMPLATE_MISPRIMING_TH=0.0
P3_COMMENT=default values of parameters using thermodynamic approach
SEQUENCE_TEMPLATE=AAGCCGAGGATGACTTCACTTTCCGACCCTGCAGAGCCGAGTTTCACCCACGAGTGAAGACTCCGCCAGCCTTGCCCCTCGGGCCTCTCCCTCAACTCACCATGATGACCGTCAGGCTGAGGAGGCAGGCGGACTGAGGCGTGGTCCGCGGGGCTCAGGGAGGTGGCTCCTGCGCCGG
PRIMER_LEFT_EXPLAIN=considered 1290, GC content failed 10, low tm 60, high tm 1039, ok 181
PRIMER_RIGHT_EXPLAIN=considered 1290, GC content failed 32, low tm 38, high tm 1096, ok 124
PRIMER_INTERNAL_EXPLAIN=considered 1565, GC content failed 32, low tm 60, high tm 1290, ok 183
PRIMER_PAIR_EXPLAIN=considered 25, unacceptable product size 20, ok 5
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=0.340791
PRIMER_LEFT_0_PENALTY=0.302821
PRIMER_RIGHT_0_PENALTY=0.037969
PRIMER_INTERNAL_0_PENALTY=0.712692
PRIMER_LEFT_0_SEQUENCE=GACTTCACTTTCCGACCCTG
PRIMER_RIGHT_0_SEQUENCE=TGACGGTCATCATGGTGAGT
PRIMER_INTERNAL_0_SEQUENCE=CACCCACGAGTGAAGACTCC
PRIMER_LEFT_0=11,20
PRIMER_RIGHT_0=113,20
PRIMER_INTERNAL_0=44,20
PRIMER_LEFT_0_TM=59.697
PRIMER_RIGHT_0_TM=59.962
PRIMER_INTERNAL_0_TM=60.713
PRIMER_LEFT_0_GC_PERCENT=55.000
PRIMER_RIGHT_0_GC_PERCENT=50.000
PRIMER_INTERNAL_0_GC_PERCENT=60.000
PRIMER_INTERNAL_0_SELF_ANY_TH=1.74
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=3.73
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=40.53
PRIMER_LEFT_0_END_STABILITY=9.7000
PRIMER_RIGHT_0_END_STABILITY=6.4000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=2.0989
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=7.9109
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=103
PRIMER_PAIR_0_PRODUCT_TM=78.8
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=7.91
PRIMER_PAIR_1_PENALTY=0.340791
PRIMER_LEFT_1_PENALTY=0.302821
PRIMER_RIGHT_1_PENALTY=0.037969
PRIMER_INTERNAL_1_PENALTY=0.712692
PRIMER_LEFT_1_SEQUENCE=TGACTTCACTTTCCGACCCT
PRIMER_RIGHT_1_SEQUENCE=TGACGGTCATCATGGTGAGT
PRIMER_INTERNAL_1_SEQUENCE=CACCCACGAGTGAAGACTCC
PRIMER_LEFT_1=10,20
PRIMER_RIGHT_1=113,20
PRIMER_INTERNAL_1=44,20
PRIMER_LEFT_1_TM=59.697
PRIMER_RIGHT_1_TM=59.962
PRIMER_INTERNAL_1_TM=60.713
PRIMER_LEFT_1_GC_PERCENT=50.000
PRIMER_RIGHT_1_GC_PERCENT=50.000
PRIMER_INTERNAL_1_GC_PERCENT=60.000
PRIMER_INTERNAL_1_SELF_ANY_TH=1.74
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=3.73
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=40.53
PRIMER_LEFT_1_END_STABILITY=9.1000
PRIMER_RIGHT_1_END_STABILITY=6.4000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=7.9109
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=104
PRIMER_PAIR_1_PRODUCT_TM=78.6
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=7.91
PRIMER_PAIR_2_PENALTY=0.415766
PRIMER_LEFT_2_PENALTY=0.302821
PRIMER_RIGHT_2_PENALTY=0.112944
PRIMER_INTERNAL_2_PENALTY=0.712692
PRIMER_LEFT_2_SEQUENCE=GACTTCACTTTCCGACCCTG
PRIMER_RIGHT_2_SEQUENCE=CTGACGGTCATCATGGTGAG
PRIMER_INTERNAL_2_SEQUENCE=CACCCACGAGTGAAGACTCC
PRIMER_LEFT_2=11,20
PRIMER_RIGHT_2=114,20
PRIMER_INTERNAL_2=44,20
PRIMER_LEFT_2_TM=59.697
PRIMER_RIGHT_2_TM=60.113
PRIMER_INTERNAL_2_TM=60.713
PRIMER_LEFT_2_GC_PERCENT=55.000
PRIMER_RIGHT_2_GC_PERCENT=55.000
PRIMER_INTERNAL_2_GC_PERCENT=60.000
PRIMER_INTERNAL_2_SELF_ANY_TH=1.74
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=3.73
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=40.53
PRIMER_LEFT_2_END_STABILITY=9.7000
PRIMER_RIGHT_2_END_STABILITY=6.4000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=2.0989
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=19.2848
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=104
PRIMER_PAIR_2_PRODUCT_TM=79.0
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=19.28
PRIMER_PAIR_3_PENALTY=0.415766
PRIMER_LEFT_3_PENALTY=0.302821
PRIMER_RIGHT_3_PENALTY=0.112944
PRIMER_INTERNAL_3_PENALTY=0.712692
PRIMER_LEFT_3_SEQUENCE=TGACTTCACTTTCCGACCCT
PRIMER_RIGHT_3_SEQUENCE=CTGACGGTCATCATGGTGAG
PRIMER_INTERNAL_3_SEQUENCE=CACCCACGAGTGAAGACTCC
PRIMER_LEFT_3=10,20
PRIMER_RIGHT_3=114,20
PRIMER_INTERNAL_3=44,20
PRIMER_LEFT_3_TM=59.697
PRIMER_RIGHT_3_TM=60.113
PRIMER_INTERNAL_3_TM=60.713
PRIMER_LEFT_3_GC_PERCENT=50.000
PRIMER_RIGHT_3_GC_PERCENT=55.000
PRIMER_INTERNAL_3_GC_PERCENT=60.000
PRIMER_INTERNAL_3_SELF_ANY_TH=1.74
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=3.73
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=40.53
PRIMER_LEFT_3_END_STABILITY=9.1000
PRIMER_RIGHT_3_END_STABILITY=6.4000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=19.2848
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=105
PRIMER_PAIR_3_PRODUCT_TM=78.8
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=19.28
PRIMER_PAIR_4_PENALTY=0.749760
PRIMER_LEFT_4_PENALTY=0.711791
PRIMER_RIGHT_4_PENALTY=0.037969
PRIMER_INTERNAL_4_PENALTY=0.712692
PRIMER_LEFT_4_SEQUENCE=GCCGAGGATGACTTCACTTT
PRIMER_RIGHT_4_SEQUENCE=TGACGGTCATCATGGTGAGT
PRIMER_INTERNAL_4_SEQUENCE=CACCCACGAGTGAAGACTCC
PRIMER_LEFT_4=2,20
PRIMER_RIGHT_4=113,20
PRIMER_INTERNAL_4=44,20
PRIMER_LEFT_4_TM=59.288
PRIMER_RIGHT_4_TM=59.962
PRIMER_INTERNAL_4_TM=60.713
PRIMER_LEFT_4_GC_PERCENT=50.000
PRIMER_RIGHT_4_GC_PERCENT=50.000
PRIMER_INTERNAL_4_GC_PERCENT=60.000
PRIMER_INTERNAL_4_SELF_ANY_TH=1.74
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=3.73
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=40.53
PRIMER_LEFT_4_END_STABILITY=6.7000
PRIMER_RIGHT_4_END_STABILITY=6.4000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=7.9109
PRIMER_PAIR_4_COMPL_ANY_TH=7.99
PRIMER_PAIR_4_COMPL_END_TH=3.92
PRIMER_PAIR_4_PRODUCT_SIZE=112
PRIMER_PAIR_4_PRODUCT_TM=79.4
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=7.91
=
SEQUENCE_ID=any_1
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_ANY_TH=10.0
PRIMER_PAIR_MAX_COMPL_ANY_TH=47.0
PRIMER_INTERNAL_MAX_SELF_ANY_TH=47.0
PRIMER_WT_SELF_ANY_TH=0.0
PRIMER_INTERNAL_WT_SELF_ANY_TH=0.0
PRIMER_PAIR_WT_COMPL_ANY_TH=0.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_LEFT_EXPLAIN=considered 3397, low tm 897, high tm 1543, long poly-x seq 22, ok 935
PRIMER_RIGHT_EXPLAIN=considered 3397, low tm 839, high tm 1673, high hairpin stability 1, long poly-x seq 22, ok 862
PRIMER_INTERNAL_EXPLAIN=considered 3672, low tm 953, high tm 1734, high hairpin stability 14, long poly-x seq 22, ok 949
PRIMER_PAIR_EXPLAIN=considered 58, unacceptable product size 51, ok 7
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=0.030625
PRIMER_LEFT_0_PENALTY=0.011300
PRIMER_RIGHT_0_PENALTY=0.019325
PRIMER_INTERNAL_0_PENALTY=1.584087
PRIMER_LEFT_0_SEQUENCE=CAAGCAATACGACCTGCTGA
PRIMER_RIGHT_0_SEQUENCE=ATCGCAAGTGAGGATGAGCT
PRIMER_INTERNAL_0_SEQUENCE=CGTGTGCAAAACATTTTCACA
PRIMER_LEFT_0=17,20
PRIMER_RIGHT_0=168,20
PRIMER_INTERNAL_0=116,21
PRIMER_LEFT_0_TM=60.011
PRIMER_RIGHT_0_TM=59.981
PRIMER_INTERNAL_0_TM=60.584
PRIMER_LEFT_0_GC_PERCENT=50.000
PRIMER_RIGHT_0_GC_PERCENT=50.000
PRIMER_INTERNAL_0_GC_PERCENT=38.095
PRIMER_INTERNAL_0_SELF_ANY_TH=8.89
PRIMER_LEFT_0_SELF_ANY_TH=6.12
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=8.89
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=42.83
PRIMER_RIGHT_0_HAIRPIN_TH=39.48
PRIMER_INTERNAL_0_HAIRPIN_TH=37.82
PRIMER_LEFT_0_END_STABILITY=8.2000
PRIMER_RIGHT_0_END_STABILITY=7.9000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=13.2447
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=21.8859
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=152
PRIMER_PAIR_0_PRODUCT_TM=71.9
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=21.89
PRIMER_PAIR_1_PENALTY=0.074511
PRIMER_LEFT_1_PENALTY=0.011300
PRIMER_RIGHT_1_PENALTY=0.063211
PRIMER_INTERNAL_1_PENALTY=0.019325
PRIMER_LEFT_1_SEQUENCE=CAAGCAATACGACCTGCTGA
PRIMER_RIGHT_1_SEQUENCE=CAAAAGATTTTGGCGGGTAA
PRIMER_INTERNAL_1_SEQUENCE=AGCTCATCCTCACTTGCGAT
PRIMER_LEFT_1=17,20
PRIMER_RIGHT_1=191,20
PRIMER_INTERNAL_1=149,20
PRIMER_LEFT_1_TM=60.011
PRIMER_RIGHT_1_TM=59.937
PRIMER_INTERNAL_1_TM=59.981
PRIMER_LEFT_1_GC_PERCENT=50.000
PRIMER_RIGHT_1_GC_PERCENT=40.000
PRIMER_INTERNAL_1_GC_PERCENT=50.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=6.12
PRIMER_RIGHT_1_SELF_ANY_TH=5.65
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=42.83
PRIMER_RIGHT_1_HAIRPIN_TH=39.80
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.2000
PRIMER_RIGHT_1_END_STABILITY=7.2000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=13.2447
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=2.5469
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=175
PRIMER_PAIR_1_PRODUCT_TM=72.4
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=15.79
PRIMER_PAIR_2_PENALTY=0.258931
PRIMER_LEFT_2_PENALTY=0.011300
PRIMER_RIGHT_2_PENALTY=0.247631
PRIMER_INTERNAL_2_PENALTY=0.019325
PRIMER_LEFT_2_SEQUENCE=CAAGCAATACGACCTGCTGA
PRIMER_RIGHT_2_SEQUENCE=CCGTACGCACTATAGGAGCC
PRIMER_INTERNAL_2_SEQUENCE=AGCTCATCCTCACTTGCGAT
PRIMER_LEFT_2=17,20
PRIMER_RIGHT_2=368,20
PRIMER_INTERNAL_2=149,20
PRIMER_LEFT_2_TM=60.011
PRIMER_RIGHT_2_TM=59.752
PRIMER_INTERNAL_2_TM=59.981
PRIMER_LEFT_2_GC_PERCENT=50.000
PRIMER_RIGHT_2_GC_PERCENT=60.000
PRIMER_INTERNAL_2_GC_PERCENT=50.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=6.12
PRIMER_RIGHT_2_SELF_ANY_TH=7.28
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=42.83
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.2000
PRIMER_RIGHT_2_END_STABILITY=9.4000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=13.2447
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=26.4047
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=352
PRIMER_PAIR_2_PRODUCT_TM=77.2
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=29.35
PRIMER_PAIR_3_PENALTY=0.266957
PRIMER_LEFT_3_PENALTY=0.019325
PRIMER_RIGHT_3_PENALTY=0.247631
PRIMER_INTERNAL_3_PENALTY=0.063211
PRIMER_LEFT_3_SEQUENCE=AGCTCATCCTCACTTGCGAT
PRIMER_RIGHT_3_SEQUENCE=CCGTACGCACTATAGGAGCC
PRIMER_INTERNAL_3_SEQUENCE=TTACCCGCCAAAATCTTTTG
PRIMER_LEFT_3=149,20
PRIMER_RIGHT_3=368,20
PRIMER_INTERNAL_3=172,20
PRIMER_LEFT_3_TM=59.981
PRIMER_RIGHT_3_TM=59.752
PRIMER_INTERNAL_3_TM=59.937
PRIMER_LEFT_3_GC_PERCENT=50.000
PRIMER_RIGHT_3_GC_PERCENT=60.000
PRIMER_INTERNAL_3_GC_PERCENT=40.000
PRIMER_INTERNAL_3_SELF_ANY_TH=4.21
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=7.28
PRIMER_INTERNAL_3_SELF_END_TH=4.21
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=43.62
PRIMER_LEFT_3_END_STABILITY=9.8000
PRIMER_RIGHT_3_END_STABILITY=9.4000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=6.0817
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=26.4047
PRIMER_PAIR_3_COMPL_ANY_TH=9.42
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=220
PRIMER_PAIR_3_PRODUCT_TM=78.4
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=32.49
PRIMER_PAIR_4_PENALTY=0.309424
PRIMER_LEFT_4_PENALTY=0.011300
PRIMER_RIGHT_4_PENALTY=0.298125
PRIMER_INTERNAL_4_PENALTY=0.019325
PRIMER_LEFT_4_SEQUENCE=CAAGCAATACGACCTGCTGA
PRIMER_RIGHT_4_SEQUENCE=CACTAAAGCGGGATTTGAGG
PRIMER_INTERNAL_4_SEQUENCE=AGCTCATCCTCACTTGCGAT
PRIMER_LEFT_4=17,20
PRIMER_RIGHT_4=232,20
PRIMER_INTERNAL_4=149,20
PRIMER_LEFT_4_TM=60.011
PRIMER_RIGHT_4_TM=59.702
PRIMER_INTERNAL_4_TM=59.981
PRIMER_LEFT_4_GC_PERCENT=50.000
PRIMER_RIGHT_4_GC_PERCENT=50.000
PRIMER_INTERNAL_4_GC_PERCENT=50.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=6.12
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=42.83
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.2000
PRIMER_RIGHT_4_END_STABILITY=8.2000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=13.2447
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=7.2923
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=216
PRIMER_PAIR_4_PRODUCT_TM=74.4
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=20.54
=
SEQUENCE_ID=any_salt
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_SALT_CORRECTIONS=1
PRIMER_SALT_MONOVALENT=50.0
PRIMER_SALT_DIVALENT=2.5
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_ANY_TH=60.0
PRIMER_PAIR_MAX_COMPL_ANY_TH=47.0
PRIMER_INTERNAL_MAX_SELF_ANY_TH=47.0
PRIMER_WT_SELF_ANY_TH=0.0
PRIMER_INTERNAL_WT_SELF_ANY_TH=0.0
PRIMER_PAIR_WT_COMPL_ANY_TH=0.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3397, low tm 52, high tm 3139, high hairpin stability 1, long poly-x seq 22, ok 183
PRIMER_RIGHT_EXPLAIN=considered 3397, low tm 59, high tm 3137, long poly-x seq 22, ok 179
PRIMER_INTERNAL_EXPLAIN=considered 3672, low tm 432, high tm 2545, high hairpin stability 10, long poly-x seq 22, ok 663
PRIMER_PAIR_EXPLAIN=considered 18, unacceptable product size 11, high template mispriming score 1, ok 6
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.177993
PRIMER_LEFT_0_PENALTY=0.842396
PRIMER_RIGHT_0_PENALTY=0.335596
PRIMER_INTERNAL_0_PENALTY=0.132562
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.227701
PRIMER_LEFT_1_PENALTY=0.842396
PRIMER_RIGHT_1_PENALTY=0.385305
PRIMER_INTERNAL_1_PENALTY=0.132562
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.422920
PRIMER_LEFT_2_PENALTY=1.087323
PRIMER_RIGHT_2_PENALTY=0.335596
PRIMER_INTERNAL_2_PENALTY=0.132562
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.437628
PRIMER_LEFT_3_PENALTY=1.102032
PRIMER_RIGHT_3_PENALTY=0.335596
PRIMER_INTERNAL_3_PENALTY=0.132562
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.472628
PRIMER_LEFT_4_PENALTY=1.087323
PRIMER_RIGHT_4_PENALTY=0.385305
PRIMER_INTERNAL_4_PENALTY=0.132562
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=any_weight
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_SALT_CORRECTIONS=1
PRIMER_SALT_MONOVALENT=50.0
PRIMER_SALT_DIVALENT=2.5
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_ANY_TH=47.0
PRIMER_PAIR_MAX_COMPL_ANY_TH=47.0
PRIMER_INTERNAL_MAX_SELF_ANY_TH=47.0
PRIMER_WT_SELF_ANY_TH=1.0
PRIMER_INTERNAL_WT_SELF_ANY_TH=0.1
PRIMER_PAIR_WT_COMPL_ANY_TH=0.7
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3397, low tm 52, high tm 3139, high hairpin stability 1, long poly-x seq 22, ok 183
PRIMER_RIGHT_EXPLAIN=considered 3397, low tm 59, high tm 3137, long poly-x seq 22, ok 179
PRIMER_INTERNAL_EXPLAIN=considered 3672, low tm 432, high tm 2545, high hairpin stability 10, long poly-x seq 22, ok 663
PRIMER_PAIR_EXPLAIN=considered 20, unacceptable product size 11, high template mispriming score 3, ok 6
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.226125
PRIMER_LEFT_0_PENALTY=0.859989
PRIMER_RIGHT_0_PENALTY=0.353561
PRIMER_INTERNAL_0_PENALTY=0.134205
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.275443
PRIMER_LEFT_1_PENALTY=0.859989
PRIMER_RIGHT_1_PENALTY=0.403040
PRIMER_INTERNAL_1_PENALTY=0.134205
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.470977
PRIMER_LEFT_2_PENALTY=1.104840
PRIMER_RIGHT_2_PENALTY=0.353561
PRIMER_INTERNAL_2_PENALTY=0.134205
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.486058
PRIMER_LEFT_3_PENALTY=1.119921
PRIMER_RIGHT_3_PENALTY=0.353561
PRIMER_INTERNAL_3_PENALTY=0.134205
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.520295
PRIMER_LEFT_4_PENALTY=1.104840
PRIMER_RIGHT_4_PENALTY=0.403040
PRIMER_INTERNAL_4_PENALTY=0.134205
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=any_2
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_SALT_CORRECTIONS=1
PRIMER_SALT_MONOVALENT=50.0
PRIMER_SALT_DIVALENT=2.5
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_ANY_TH=5.0
PRIMER_PAIR_MAX_COMPL_ANY_TH=5.0
PRIMER_INTERNAL_MAX_SELF_ANY_TH=5.0
PRIMER_WT_SELF_ANY_TH=1.0
PRIMER_INTERNAL_WT_SELF_ANY_TH=1.0
PRIMER_PAIR_WT_COMPL_ANY_TH=1.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3146, low tm 52, high tm 2915, high any compl 33, high hairpin stability 1, long poly-x seq 22, ok 123
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3126, low tm 432, high tm 2140, high any compl 83, high hairpin stability 10, long poly-x seq 22, ok 439
PRIMER_PAIR_EXPLAIN=considered 23, unacceptable product size 12, high template mispriming score 3, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.231515
PRIMER_LEFT_0_PENALTY=0.859989
PRIMER_RIGHT_0_PENALTY=0.353561
PRIMER_INTERNAL_0_PENALTY=0.148991
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.280764
PRIMER_LEFT_1_PENALTY=0.859989
PRIMER_RIGHT_1_PENALTY=0.403040
PRIMER_INTERNAL_1_PENALTY=0.148991
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.476366
PRIMER_LEFT_2_PENALTY=1.104840
PRIMER_RIGHT_2_PENALTY=0.353561
PRIMER_INTERNAL_2_PENALTY=0.148991
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.491447
PRIMER_LEFT_3_PENALTY=1.119921
PRIMER_RIGHT_3_PENALTY=0.353561
PRIMER_INTERNAL_3_PENALTY=0.148991
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.525615
PRIMER_LEFT_4_PENALTY=1.104840
PRIMER_RIGHT_4_PENALTY=0.403040
PRIMER_INTERNAL_4_PENALTY=0.148991
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=any_no_effect
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_SALT_CORRECTIONS=1
PRIMER_SALT_MONOVALENT=50.0
PRIMER_SALT_DIVALENT=2.5
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_ANY_TH=0.0
PRIMER_PAIR_MAX_COMPL_ANY_TH=0.0
PRIMER_INTERNAL_MAX_SELF_ANY_TH=0.0
PRIMER_WT_SELF_ANY_TH=0.0
PRIMER_INTERNAL_WT_SELF_ANY_TH=0.0
PRIMER_PAIR_WT_COMPL_ANY_TH=0.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3397, low tm 52, high tm 3139, high hairpin stability 1, long poly-x seq 22, ok 183
PRIMER_RIGHT_EXPLAIN=considered 3397, low tm 59, high tm 3137, long poly-x seq 22, ok 179
PRIMER_INTERNAL_EXPLAIN=considered 3672, low tm 432, high tm 2545, high any compl 2, high hairpin stability 10, long poly-x seq 22, ok 661
PRIMER_PAIR_EXPLAIN=considered 18, unacceptable product size 11, high template mispriming score 1, ok 6
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.177993
PRIMER_LEFT_0_PENALTY=0.842396
PRIMER_RIGHT_0_PENALTY=0.335596
PRIMER_INTERNAL_0_PENALTY=0.132562
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.227701
PRIMER_LEFT_1_PENALTY=0.842396
PRIMER_RIGHT_1_PENALTY=0.385305
PRIMER_INTERNAL_1_PENALTY=0.132562
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.422920
PRIMER_LEFT_2_PENALTY=1.087323
PRIMER_RIGHT_2_PENALTY=0.335596
PRIMER_INTERNAL_2_PENALTY=0.132562
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.437628
PRIMER_LEFT_3_PENALTY=1.102032
PRIMER_RIGHT_3_PENALTY=0.335596
PRIMER_INTERNAL_3_PENALTY=0.132562
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.472628
PRIMER_LEFT_4_PENALTY=1.087323
PRIMER_RIGHT_4_PENALTY=0.385305
PRIMER_INTERNAL_4_PENALTY=0.132562
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=end_th1
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_END_TH=10.0
PRIMER_PAIR_MAX_COMPL_END_TH=10.0
PRIMER_INTERNAL_MAX_SELF_END_TH=10.0
PRIMER_WT_SELF_END_TH=0.0
PRIMER_INTERNAL_WT_SELF_END_TH=0.0
PRIMER_PAIR_WT_COMPL_END_TH=0.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3397, low tm 52, high tm 3139, high hairpin stability 1, long poly-x seq 22, ok 183
PRIMER_RIGHT_EXPLAIN=considered 3397, low tm 59, high tm 3137, long poly-x seq 22, ok 179
PRIMER_INTERNAL_EXPLAIN=considered 3672, low tm 432, high tm 2545, high any compl 2, high hairpin stability 10, long poly-x seq 22, ok 661
PRIMER_PAIR_EXPLAIN=considered 18, unacceptable product size 11, high template mispriming score 1, ok 6
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.177993
PRIMER_LEFT_0_PENALTY=0.842396
PRIMER_RIGHT_0_PENALTY=0.335596
PRIMER_INTERNAL_0_PENALTY=0.132562
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.227701
PRIMER_LEFT_1_PENALTY=0.842396
PRIMER_RIGHT_1_PENALTY=0.385305
PRIMER_INTERNAL_1_PENALTY=0.132562
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.422920
PRIMER_LEFT_2_PENALTY=1.087323
PRIMER_RIGHT_2_PENALTY=0.335596
PRIMER_INTERNAL_2_PENALTY=0.132562
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.437628
PRIMER_LEFT_3_PENALTY=1.102032
PRIMER_RIGHT_3_PENALTY=0.335596
PRIMER_INTERNAL_3_PENALTY=0.132562
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.472628
PRIMER_LEFT_4_PENALTY=1.087323
PRIMER_RIGHT_4_PENALTY=0.385305
PRIMER_INTERNAL_4_PENALTY=0.132562
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=end_th2
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_SELF_END_TH=47.0
PRIMER_PAIR_MAX_COMPL_END_TH=47.0
PRIMER_INTERNAL_MAX_SELF_END_TH=47.0
PRIMER_WT_SELF_END_TH=0.1
PRIMER_INTERNAL_WT_SELF_END_TH=0.01
PRIMER_PAIR_WT_COMPL_END_TH=1.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 1, long poly-x seq 22, ok 111
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 10, long poly-x seq 22, ok 376
PRIMER_PAIR_EXPLAIN=considered 23, unacceptable product size 12, high template mispriming score 3, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.199513
PRIMER_LEFT_0_PENALTY=0.844155
PRIMER_RIGHT_0_PENALTY=0.337393
PRIMER_INTERNAL_0_PENALTY=0.132726
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.248969
PRIMER_LEFT_1_PENALTY=0.844155
PRIMER_RIGHT_1_PENALTY=0.387078
PRIMER_INTERNAL_1_PENALTY=0.132726
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.444433
PRIMER_LEFT_2_PENALTY=1.089075
PRIMER_RIGHT_2_PENALTY=0.337393
PRIMER_INTERNAL_2_PENALTY=0.132726
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.459178
PRIMER_LEFT_3_PENALTY=1.103821
PRIMER_RIGHT_3_PENALTY=0.337393
PRIMER_INTERNAL_3_PENALTY=0.132726
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.493888
PRIMER_LEFT_4_PENALTY=1.089075
PRIMER_RIGHT_4_PENALTY=0.387078
PRIMER_INTERNAL_4_PENALTY=0.132726
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=hairpin1
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_HAIRPIN_TH=50.0
PRIMER_INTERNAL_MAX_HAIRPIN_TH=50.0
PRIMER_WT_HAIRPIN_TH=1.0
PRIMER_INTERNAL_WT_HAIRPIN_TH=1.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 8, long poly-x seq 22, ok 104
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 10, long poly-x seq 22, ok 376
PRIMER_PAIR_EXPLAIN=considered 19, unacceptable product size 8, high template mispriming score 3, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.235070
PRIMER_LEFT_0_PENALTY=0.861748
PRIMER_RIGHT_0_PENALTY=0.355358
PRIMER_INTERNAL_0_PENALTY=0.149156
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.284296
PRIMER_LEFT_1_PENALTY=0.861748
PRIMER_RIGHT_1_PENALTY=0.404813
PRIMER_INTERNAL_1_PENALTY=0.149156
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.479914
PRIMER_LEFT_2_PENALTY=1.106592
PRIMER_RIGHT_2_PENALTY=0.355358
PRIMER_INTERNAL_2_PENALTY=0.149156
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.495033
PRIMER_LEFT_3_PENALTY=1.121710
PRIMER_RIGHT_3_PENALTY=0.355358
PRIMER_INTERNAL_3_PENALTY=0.149156
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.529140
PRIMER_LEFT_4_PENALTY=1.106592
PRIMER_RIGHT_4_PENALTY=0.404813
PRIMER_INTERNAL_4_PENALTY=0.149156
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=hairpin2
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_HAIRPIN_TH=40.0
PRIMER_INTERNAL_MAX_HAIRPIN_TH=40.0
PRIMER_WT_HAIRPIN_TH=0.0
PRIMER_INTERNAL_WT_HAIRPIN_TH=0.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 1, long poly-x seq 22, ok 111
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 10, long poly-x seq 22, ok 376
PRIMER_PAIR_EXPLAIN=considered 23, unacceptable product size 12, high template mispriming score 3, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.199513
PRIMER_LEFT_0_PENALTY=0.844155
PRIMER_RIGHT_0_PENALTY=0.337393
PRIMER_INTERNAL_0_PENALTY=0.132726
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.248969
PRIMER_LEFT_1_PENALTY=0.844155
PRIMER_RIGHT_1_PENALTY=0.387078
PRIMER_INTERNAL_1_PENALTY=0.132726
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.444433
PRIMER_LEFT_2_PENALTY=1.089075
PRIMER_RIGHT_2_PENALTY=0.337393
PRIMER_INTERNAL_2_PENALTY=0.132726
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.459178
PRIMER_LEFT_3_PENALTY=1.103821
PRIMER_RIGHT_3_PENALTY=0.337393
PRIMER_INTERNAL_3_PENALTY=0.132726
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.493888
PRIMER_LEFT_4_PENALTY=1.089075
PRIMER_RIGHT_4_PENALTY=0.387078
PRIMER_INTERNAL_4_PENALTY=0.132726
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=hairpin3
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_HAIRPIN_TH=0.0
PRIMER_INTERNAL_MAX_HAIRPIN_TH=47.0
PRIMER_PICK_ANYWAY=1
PRIMER_WT_HAIRPIN_TH=0.0
PRIMER_INTERNAL_WT_HAIRPIN_TH=1.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 1, long poly-x seq 22, ok 111
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 28, long poly-x seq 22, ok 358
PRIMER_PAIR_EXPLAIN=considered 23, unacceptable product size 12, high template mispriming score 3, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.199513
PRIMER_LEFT_0_PENALTY=0.844155
PRIMER_RIGHT_0_PENALTY=0.337393
PRIMER_INTERNAL_0_PENALTY=0.149156
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.248969
PRIMER_LEFT_1_PENALTY=0.844155
PRIMER_RIGHT_1_PENALTY=0.387078
PRIMER_INTERNAL_1_PENALTY=0.149156
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.444433
PRIMER_LEFT_2_PENALTY=1.089075
PRIMER_RIGHT_2_PENALTY=0.337393
PRIMER_INTERNAL_2_PENALTY=0.149156
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.459178
PRIMER_LEFT_3_PENALTY=1.103821
PRIMER_RIGHT_3_PENALTY=0.337393
PRIMER_INTERNAL_3_PENALTY=0.149156
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.493888
PRIMER_LEFT_4_PENALTY=1.089075
PRIMER_RIGHT_4_PENALTY=0.387078
PRIMER_INTERNAL_4_PENALTY=0.149156
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=template1
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_MAX_TEMPLATE_MISPRIMING_TH=45.0
PRIMER_WT_TEMPLATE_MISPRIMING_TH=1.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 1, long poly-x seq 22, ok 111
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 28, long poly-x seq 22, ok 358
PRIMER_PAIR_EXPLAIN=considered 20, unacceptable product size 11, high template mispriming score 1, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.243737
PRIMER_LEFT_0_PENALTY=0.861748
PRIMER_RIGHT_0_PENALTY=0.364025
PRIMER_INTERNAL_0_PENALTY=0.149156
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.289853
PRIMER_LEFT_1_PENALTY=0.861748
PRIMER_RIGHT_1_PENALTY=0.410370
PRIMER_INTERNAL_1_PENALTY=0.149156
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.488581
PRIMER_LEFT_2_PENALTY=1.106592
PRIMER_RIGHT_2_PENALTY=0.364025
PRIMER_INTERNAL_2_PENALTY=0.149156
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.504502
PRIMER_LEFT_3_PENALTY=1.122512
PRIMER_RIGHT_3_PENALTY=0.364025
PRIMER_INTERNAL_3_PENALTY=0.149156
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.534697
PRIMER_LEFT_4_PENALTY=1.106592
PRIMER_RIGHT_4_PENALTY=0.410370
PRIMER_INTERNAL_4_PENALTY=0.149156
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=template2
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_PAIR_MAX_TEMPLATE_MISPRIMING_TH=50.0
PRIMER_PAIR_WT_TEMPLATE_MISPRIMING_TH=1.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 1, long poly-x seq 22, ok 111
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 28, long poly-x seq 22, ok 358
PRIMER_PAIR_EXPLAIN=considered 23, unacceptable product size 12, high template mispriming score 3, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.270369
PRIMER_LEFT_0_PENALTY=0.861748
PRIMER_RIGHT_0_PENALTY=0.364025
PRIMER_INTERNAL_0_PENALTY=0.149156
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.313145
PRIMER_LEFT_1_PENALTY=0.861748
PRIMER_RIGHT_1_PENALTY=0.410370
PRIMER_INTERNAL_1_PENALTY=0.149156
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.515213
PRIMER_LEFT_2_PENALTY=1.106592
PRIMER_RIGHT_2_PENALTY=0.364025
PRIMER_INTERNAL_2_PENALTY=0.149156
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.532950
PRIMER_LEFT_3_PENALTY=1.122512
PRIMER_RIGHT_3_PENALTY=0.364025
PRIMER_INTERNAL_3_PENALTY=0.149156
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.557989
PRIMER_LEFT_4_PENALTY=1.106592
PRIMER_RIGHT_4_PENALTY=0.410370
PRIMER_INTERNAL_4_PENALTY=0.149156
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=template2_and_repeat_library
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_MISPRIMING_LIBRARY=../src/humrep_and_simple.txt
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=1
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_PAIR_MAX_TEMPLATE_MISPRIMING_TH=50.0
PRIMER_PAIR_WT_TEMPLATE_MISPRIMING_TH=1.0
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 1, long poly-x seq 22, ok 111
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, long poly-x seq 18, ok 142
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 28, long poly-x seq 22, ok 358
PRIMER_PAIR_EXPLAIN=considered 23, unacceptable product size 12, high template mispriming score 3, ok 8
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=1.270369
PRIMER_LEFT_0_PENALTY=0.861748
PRIMER_RIGHT_0_PENALTY=0.364025
PRIMER_INTERNAL_0_PENALTY=0.149156
PRIMER_LEFT_0_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_0_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_0_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_0=98,20
PRIMER_RIGHT_0=254,20
PRIMER_INTERNAL_0=145,20
PRIMER_LEFT_0_TM=60.842
PRIMER_RIGHT_0_TM=59.664
PRIMER_INTERNAL_0_TM=59.867
PRIMER_LEFT_0_GC_PERCENT=30.000
PRIMER_RIGHT_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_GC_PERCENT=45.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_LIBRARY_MISPRIMING=12.00, MER33 Nonautonomous DNA transposon
PRIMER_RIGHT_0_LIBRARY_MISPRIMING=11.00, reverse Tigger2 Autonomous DNA transposon
PRIMER_PAIR_0_LIBRARY_MISPRIMING=20.00, reverse Tigger2 Autonomous DNA transposon
PRIMER_LEFT_0_END_STABILITY=8.7000
PRIMER_RIGHT_0_END_STABILITY=5.1000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=157
PRIMER_PAIR_0_PRODUCT_TM=84.6
PRIMER_PAIR_0_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_1_PENALTY=1.313145
PRIMER_LEFT_1_PENALTY=0.861748
PRIMER_RIGHT_1_PENALTY=0.410370
PRIMER_INTERNAL_1_PENALTY=0.149156
PRIMER_LEFT_1_SEQUENCE=ATTTTTCTATCTGTACAACG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=98,20
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=60.842
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=30.000
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_LIBRARY_MISPRIMING=12.00, MER33 Nonautonomous DNA transposon
PRIMER_RIGHT_1_LIBRARY_MISPRIMING=12.00, MER41 Medium reiteration sequence MER41
PRIMER_PAIR_1_LIBRARY_MISPRIMING=19.00, MIR mammalian-wide interspersed repeat (MIR) - a consensus
PRIMER_LEFT_1_END_STABILITY=8.7000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=146
PRIMER_PAIR_1_PRODUCT_TM=84.5
PRIMER_PAIR_1_TEMPLATE_MISPRIMING_TH=13.45
PRIMER_PAIR_2_PENALTY=1.515213
PRIMER_LEFT_2_PENALTY=1.106592
PRIMER_RIGHT_2_PENALTY=0.364025
PRIMER_INTERNAL_2_PENALTY=0.149156
PRIMER_LEFT_2_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_2_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=99,20
PRIMER_RIGHT_2=254,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=61.087
PRIMER_RIGHT_2_TM=59.664
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=30.000
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_LIBRARY_MISPRIMING=11.00, MER33 Nonautonomous DNA transposon
PRIMER_RIGHT_2_LIBRARY_MISPRIMING=11.00, reverse Tigger2 Autonomous DNA transposon
PRIMER_PAIR_2_LIBRARY_MISPRIMING=19.00, reverse L1MA9 3'-end of L1 repeat (subfamily L1MA9) - a consensus sequence
PRIMER_LEFT_2_END_STABILITY=8.1000
PRIMER_RIGHT_2_END_STABILITY=5.1000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=156
PRIMER_PAIR_2_PRODUCT_TM=84.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING_TH=18.12
PRIMER_PAIR_3_PENALTY=1.532950
PRIMER_LEFT_3_PENALTY=1.122512
PRIMER_RIGHT_3_PENALTY=0.364025
PRIMER_INTERNAL_3_PENALTY=0.149156
PRIMER_LEFT_3_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_3_SEQUENCE=GACCTGTAATAGCTCACTAC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=103,19
PRIMER_RIGHT_3=254,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=59.898
PRIMER_RIGHT_3_TM=59.664
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=42.105
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_LIBRARY_MISPRIMING=11.00, MER11A a consensus
PRIMER_RIGHT_3_LIBRARY_MISPRIMING=11.00, reverse Tigger2 Autonomous DNA transposon
PRIMER_PAIR_3_LIBRARY_MISPRIMING=20.00, reverse L1MA9 3'-end of L1 repeat (subfamily L1MA9) - a consensus sequence
PRIMER_LEFT_3_END_STABILITY=6.4000
PRIMER_RIGHT_3_END_STABILITY=5.1000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING_TH=2.3979
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING_TH=18.1151
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=152
PRIMER_PAIR_3_PRODUCT_TM=85.1
PRIMER_PAIR_3_TEMPLATE_MISPRIMING_TH=20.51
PRIMER_PAIR_4_PENALTY=1.557989
PRIMER_LEFT_4_PENALTY=1.106592
PRIMER_RIGHT_4_PENALTY=0.410370
PRIMER_INTERNAL_4_PENALTY=0.149156
PRIMER_LEFT_4_SEQUENCE=TTTTTCTATCTGTACAACGT
PRIMER_RIGHT_4_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=99,20
PRIMER_RIGHT_4=243,20
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=61.087
PRIMER_RIGHT_4_TM=60.385
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=30.000
PRIMER_RIGHT_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_LIBRARY_MISPRIMING=11.00, MER33 Nonautonomous DNA transposon
PRIMER_RIGHT_4_LIBRARY_MISPRIMING=12.00, MER41 Medium reiteration sequence MER41
PRIMER_PAIR_4_LIBRARY_MISPRIMING=19.00, reverse L1 Human L1 interspersed repetitive sequence - full length copy
PRIMER_LEFT_4_END_STABILITY=8.1000
PRIMER_RIGHT_4_END_STABILITY=8.5000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING_TH=0.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING_TH=13.4516
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=145
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING_TH=13.45
=
SEQUENCE_ID=template3
PRIMER_TASK=pick_pcr_primers_and_hyb_probe
PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1
PRIMER_THERMODYNAMIC_TEMPLATE_ALIGNMENT=0
PRIMER_EXPLAIN_FLAG=1
PRIMER_NUM_RETURN=5
PRIMER_PRODUCT_SIZE_RANGE=50-500
PRIMER_PAIR_MAX_TEMPLATE_MISPRIMING=20
PRIMER_PAIR_WT_TEMPLATE_MISPRIMING=100
SEQUENCE_TEMPLATE=AGTGAGCTCATTCAATACAAGCAATACGACCTGCTGATTATCGACGATATTCAGTTCATCAAAGGAAAGTGAGCTCATTAAGACCGTACGATGGAAGAATTTTTCTATCTGTACAACGTGTGCAAAACATTTTCACAATGAGAAAAAACAGCTCATCCTCACTTGCGATGTTTTACCCGCCAAAATCTTTTGCACCGAAGGTATGGACGACCGCCTCAAATCCCGCTTTAGTGTAGTAGTGAGCTATTACAGGTCGTGGGGGCTGACCGCAGTAAGCTGCGGCTTTGGAACTCGAGCCGCCCGAATTGGAAATGCGCAAAACCGTGTCTATCGCCATTTTGCAGAAAAAGGCTCCTATAGTGCGTACGGAAGCGGCGGGGTACGCACTATAGACATCAGTATCG
PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC
PRIMER_LEFT_EXPLAIN=considered 3089, low tm 52, high tm 2863, high any compl 40, high hairpin stability 25, high repeat similarity 3, long poly-x seq 22, ok 84
PRIMER_RIGHT_EXPLAIN=considered 3209, low tm 59, high tm 2966, high any compl 24, high hairpin stability 38, long poly-x seq 18, ok 104
PRIMER_INTERNAL_EXPLAIN=considered 3030, low tm 432, high tm 2088, high any compl 102, high hairpin stability 28, long poly-x seq 22, ok 358
PRIMER_PAIR_EXPLAIN=considered 8911, unacceptable product size 3799, high any compl 86, no internal oligo 571, high template mispriming score 119, ok 4336
PRIMER_LEFT_NUM_RETURNED=5
PRIMER_RIGHT_NUM_RETURNED=5
PRIMER_INTERNAL_NUM_RETURNED=5
PRIMER_PAIR_NUM_RETURNED=5
PRIMER_PAIR_0_PENALTY=906.570516
PRIMER_LEFT_0_PENALTY=3.538806
PRIMER_RIGHT_0_PENALTY=3.014170
PRIMER_INTERNAL_0_PENALTY=0.220616
PRIMER_LEFT_0_SEQUENCE=TTCAATACAAGCAATACG
PRIMER_RIGHT_0_SEQUENCE=GCACACGTTGTACAGATA
PRIMER_INTERNAL_0_SEQUENCE=TCATCAAAGGAAAGTGAGCT
PRIMER_LEFT_0=10,18
PRIMER_RIGHT_0=122,18
PRIMER_INTERNAL_0=55,20
PRIMER_LEFT_0_TM=61.537
PRIMER_RIGHT_0_TM=61.012
PRIMER_INTERNAL_0_TM=59.796
PRIMER_LEFT_0_GC_PERCENT=33.333
PRIMER_RIGHT_0_GC_PERCENT=44.444
PRIMER_INTERNAL_0_GC_PERCENT=40.000
PRIMER_INTERNAL_0_SELF_ANY_TH=0.00
PRIMER_LEFT_0_SELF_ANY_TH=0.00
PRIMER_RIGHT_0_SELF_ANY_TH=0.00
PRIMER_INTERNAL_0_SELF_END_TH=0.00
PRIMER_LEFT_0_SELF_END_TH=0.00
PRIMER_RIGHT_0_SELF_END_TH=0.00
PRIMER_LEFT_0_HAIRPIN_TH=0.00
PRIMER_RIGHT_0_HAIRPIN_TH=0.00
PRIMER_INTERNAL_0_HAIRPIN_TH=0.00
PRIMER_LEFT_0_LIBRARY_MISPRIMING=10.00, TACAA Primate simple repeat pattern
PRIMER_RIGHT_0_LIBRARY_MISPRIMING=11.00, L1MA10 3'-end of L1 repeat (subfamily L1MA10) - a consensus sequence
PRIMER_PAIR_0_LIBRARY_MISPRIMING=18.00, L1 Human L1 interspersed repetitive sequence - full length copy
PRIMER_LEFT_0_END_STABILITY=7.3000
PRIMER_RIGHT_0_END_STABILITY=5.6000
PRIMER_LEFT_0_TEMPLATE_MISPRIMING=5.0000
PRIMER_RIGHT_0_TEMPLATE_MISPRIMING=5.0000
PRIMER_PAIR_0_COMPL_ANY_TH=0.00
PRIMER_PAIR_0_COMPL_END_TH=0.00
PRIMER_PAIR_0_PRODUCT_SIZE=113
PRIMER_PAIR_0_PRODUCT_TM=81.9
PRIMER_PAIR_0_TEMPLATE_MISPRIMING=9.00
PRIMER_PAIR_1_PENALTY=1001.508789
PRIMER_LEFT_1_PENALTY=1.103821
PRIMER_RIGHT_1_PENALTY=0.387078
PRIMER_INTERNAL_1_PENALTY=0.149156
PRIMER_LEFT_1_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_1_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_1_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_1=103,19
PRIMER_RIGHT_1=243,20
PRIMER_INTERNAL_1=145,20
PRIMER_LEFT_1_TM=59.898
PRIMER_RIGHT_1_TM=60.385
PRIMER_INTERNAL_1_TM=59.867
PRIMER_LEFT_1_GC_PERCENT=42.105
PRIMER_RIGHT_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_GC_PERCENT=45.000
PRIMER_INTERNAL_1_SELF_ANY_TH=0.00
PRIMER_LEFT_1_SELF_ANY_TH=0.00
PRIMER_RIGHT_1_SELF_ANY_TH=0.00
PRIMER_INTERNAL_1_SELF_END_TH=0.00
PRIMER_LEFT_1_SELF_END_TH=0.00
PRIMER_RIGHT_1_SELF_END_TH=0.00
PRIMER_LEFT_1_HAIRPIN_TH=0.00
PRIMER_RIGHT_1_HAIRPIN_TH=0.00
PRIMER_INTERNAL_1_HAIRPIN_TH=0.00
PRIMER_LEFT_1_LIBRARY_MISPRIMING=11.00, MER11A a consensus
PRIMER_RIGHT_1_LIBRARY_MISPRIMING=12.00, MER41 Medium reiteration sequence MER41
PRIMER_PAIR_1_LIBRARY_MISPRIMING=19.00, L1MA10 3'-end of L1 repeat (subfamily L1MA10) - a consensus sequence
PRIMER_LEFT_1_END_STABILITY=6.4000
PRIMER_RIGHT_1_END_STABILITY=8.5000
PRIMER_LEFT_1_TEMPLATE_MISPRIMING=5.0000
PRIMER_RIGHT_1_TEMPLATE_MISPRIMING=5.0000
PRIMER_PAIR_1_COMPL_ANY_TH=0.00
PRIMER_PAIR_1_COMPL_END_TH=0.00
PRIMER_PAIR_1_PRODUCT_SIZE=141
PRIMER_PAIR_1_PRODUCT_TM=85.0
PRIMER_PAIR_1_TEMPLATE_MISPRIMING=10.00
PRIMER_PAIR_2_PENALTY=1001.568197
PRIMER_LEFT_2_PENALTY=1.163210
PRIMER_RIGHT_2_PENALTY=0.387078
PRIMER_INTERNAL_2_PENALTY=0.149156
PRIMER_LEFT_2_SEQUENCE=CATTCAATACAAGCAATAC
PRIMER_RIGHT_2_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_2_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_2=8,19
PRIMER_RIGHT_2=243,20
PRIMER_INTERNAL_2=145,20
PRIMER_LEFT_2_TM=59.839
PRIMER_RIGHT_2_TM=60.385
PRIMER_INTERNAL_2_TM=59.867
PRIMER_LEFT_2_GC_PERCENT=31.579
PRIMER_RIGHT_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_GC_PERCENT=45.000
PRIMER_INTERNAL_2_SELF_ANY_TH=0.00
PRIMER_LEFT_2_SELF_ANY_TH=0.00
PRIMER_RIGHT_2_SELF_ANY_TH=0.00
PRIMER_INTERNAL_2_SELF_END_TH=0.00
PRIMER_LEFT_2_SELF_END_TH=0.00
PRIMER_RIGHT_2_SELF_END_TH=0.00
PRIMER_LEFT_2_HAIRPIN_TH=0.00
PRIMER_RIGHT_2_HAIRPIN_TH=0.00
PRIMER_INTERNAL_2_HAIRPIN_TH=0.00
PRIMER_LEFT_2_LIBRARY_MISPRIMING=12.00, TACAA Primate simple repeat pattern
PRIMER_RIGHT_2_LIBRARY_MISPRIMING=12.00, MER41 Medium reiteration sequence MER41
PRIMER_PAIR_2_LIBRARY_MISPRIMING=19.00, L1 Human L1 interspersed repetitive sequence - full length copy
PRIMER_LEFT_2_END_STABILITY=5.6000
PRIMER_RIGHT_2_END_STABILITY=8.5000
PRIMER_LEFT_2_TEMPLATE_MISPRIMING=5.0000
PRIMER_RIGHT_2_TEMPLATE_MISPRIMING=5.0000
PRIMER_PAIR_2_COMPL_ANY_TH=0.00
PRIMER_PAIR_2_COMPL_END_TH=0.00
PRIMER_PAIR_2_PRODUCT_SIZE=236
PRIMER_PAIR_2_PRODUCT_TM=85.7
PRIMER_PAIR_2_TEMPLATE_MISPRIMING=10.00
PRIMER_PAIR_3_PENALTY=1002.231568
PRIMER_LEFT_3_PENALTY=1.826755
PRIMER_RIGHT_3_PENALTY=0.387078
PRIMER_INTERNAL_3_PENALTY=0.149156
PRIMER_LEFT_3_SEQUENCE=ATATTCAGTTCATCAAAGG
PRIMER_RIGHT_3_SEQUENCE=GCTCACTACTACACTAAAGC
PRIMER_INTERNAL_3_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_3=46,19
PRIMER_RIGHT_3=243,20
PRIMER_INTERNAL_3=145,20
PRIMER_LEFT_3_TM=60.825
PRIMER_RIGHT_3_TM=60.385
PRIMER_INTERNAL_3_TM=59.867
PRIMER_LEFT_3_GC_PERCENT=31.579
PRIMER_RIGHT_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_GC_PERCENT=45.000
PRIMER_INTERNAL_3_SELF_ANY_TH=0.00
PRIMER_LEFT_3_SELF_ANY_TH=0.00
PRIMER_RIGHT_3_SELF_ANY_TH=0.00
PRIMER_INTERNAL_3_SELF_END_TH=0.00
PRIMER_LEFT_3_SELF_END_TH=0.00
PRIMER_RIGHT_3_SELF_END_TH=0.00
PRIMER_LEFT_3_HAIRPIN_TH=0.00
PRIMER_RIGHT_3_HAIRPIN_TH=0.00
PRIMER_INTERNAL_3_HAIRPIN_TH=0.00
PRIMER_LEFT_3_LIBRARY_MISPRIMING=10.00, THE1BR THE-1B Mammalian LTR retrotransposon internal sequence - a consensus
PRIMER_RIGHT_3_LIBRARY_MISPRIMING=12.00, MER41 Medium reiteration sequence MER41
PRIMER_PAIR_3_LIBRARY_MISPRIMING=19.00, THE1BR THE-1B Mammalian LTR retrotransposon internal sequence - a consensus
PRIMER_LEFT_3_END_STABILITY=8.5000
PRIMER_RIGHT_3_END_STABILITY=8.5000
PRIMER_LEFT_3_TEMPLATE_MISPRIMING=5.0000
PRIMER_RIGHT_3_TEMPLATE_MISPRIMING=5.0000
PRIMER_PAIR_3_COMPL_ANY_TH=0.00
PRIMER_PAIR_3_COMPL_END_TH=0.00
PRIMER_PAIR_3_PRODUCT_SIZE=198
PRIMER_PAIR_3_PRODUCT_TM=85.2
PRIMER_PAIR_3_TEMPLATE_MISPRIMING=10.00
PRIMER_PAIR_4_PENALTY=1002.371294
PRIMER_LEFT_4_PENALTY=1.103821
PRIMER_RIGHT_4_PENALTY=1.249537
PRIMER_INTERNAL_4_PENALTY=0.149156
PRIMER_LEFT_4_SEQUENCE=TCTATCTGTACAACGTGTG
PRIMER_RIGHT_4_SEQUENCE=TCACTACTACACTAAAGCG
PRIMER_INTERNAL_4_SEQUENCE=AAACAGCTCATCCTCACTTG
PRIMER_LEFT_4=103,19
PRIMER_RIGHT_4=241,19
PRIMER_INTERNAL_4=145,20
PRIMER_LEFT_4_TM=59.898
PRIMER_RIGHT_4_TM=59.752
PRIMER_INTERNAL_4_TM=59.867
PRIMER_LEFT_4_GC_PERCENT=42.105
PRIMER_RIGHT_4_GC_PERCENT=42.105
PRIMER_INTERNAL_4_GC_PERCENT=45.000
PRIMER_INTERNAL_4_SELF_ANY_TH=0.00
PRIMER_LEFT_4_SELF_ANY_TH=0.00
PRIMER_RIGHT_4_SELF_ANY_TH=0.00
PRIMER_INTERNAL_4_SELF_END_TH=0.00
PRIMER_LEFT_4_SELF_END_TH=0.00
PRIMER_RIGHT_4_SELF_END_TH=0.00
PRIMER_LEFT_4_HAIRPIN_TH=0.00
PRIMER_RIGHT_4_HAIRPIN_TH=0.00
PRIMER_INTERNAL_4_HAIRPIN_TH=0.00
PRIMER_LEFT_4_LIBRARY_MISPRIMING=11.00, MER11A a consensus
PRIMER_RIGHT_4_LIBRARY_MISPRIMING=11.00, MER41 Medium reiteration sequence MER41
PRIMER_PAIR_4_LIBRARY_MISPRIMING=19.00, L1MA10 3'-end of L1 repeat (subfamily L1MA10) - a consensus sequence
PRIMER_LEFT_4_END_STABILITY=6.4000
PRIMER_RIGHT_4_END_STABILITY=10.2000
PRIMER_LEFT_4_TEMPLATE_MISPRIMING=5.0000
PRIMER_RIGHT_4_TEMPLATE_MISPRIMING=5.0000
PRIMER_PAIR_4_COMPL_ANY_TH=0.00
PRIMER_PAIR_4_COMPL_END_TH=0.00
PRIMER_PAIR_4_PRODUCT_SIZE=139
PRIMER_PAIR_4_PRODUCT_TM=84.6
PRIMER_PAIR_4_TEMPLATE_MISPRIMING=10.00
=
