diff --git a/src/libprimer3.cc b/src/libprimer3.cc index a90ff016..ce4f10f1 100644 --- a/src/libprimer3.cc +++ b/src/libprimer3.cc @@ -7207,6 +7207,10 @@ p3_read_line(FILE *file) if ((n = strchr(p, '\n')) != NULL) { *n = '\0'; + n--; + if (*n == '\r') { + *n = '\0'; + } return s; } diff --git a/test/p3test.pl b/test/p3test.pl index 96c2c5bd..369fc41c 100755 --- a/test/p3test.pl +++ b/test/p3test.pl @@ -314,6 +314,8 @@ () 'primer_new_tasks', 'primer_new_tasks_formatted', + 'primer_windows_newlines', + 'primer_must_overlap_point', 'primer_overlap_junction', diff --git a/test/primer_windows_newlines_input b/test/primer_windows_newlines_input new file mode 100755 index 00000000..575c2abb --- /dev/null +++ b/test/primer_windows_newlines_input @@ -0,0 +1,52 @@ +PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1 +PRIMER_TM_FORMULA=1 +PRIMER_SALT_CORRECTIONS=1 +PRIMER_SALT_DIVALENT=1.5 +PRIMER_MAX_SELF_ANY=12 +PRIMER_PAIR_MAX_COMPL_ANY=12 +PRIMER_MAX_POLY_X=100 +PRIMER_MAX_SELF_END=8 +PRIMER_PAIR_MAX_COMPL_END=8 +P3_FILE_FLAG=1 +PRIMER_EXPLAIN_FLAG=1 +PRIMER_TASK=pick_pcr_primers +PRIMER_DNTP_CONC=0.0 +SEQUENCE_ID=pcr_primers +SEQUENCE_TEMPLATE=TCCTACAGCTGTGGGAAAATGatgcgtacgatccatgctagctagctactatcgattagctatcatctactatcatctactatctactacacatctactatcatctacacacacacacacactaggccgctatatcatctagcGCCGCCATGTACACACTG +SEQUENCE_TARGET=21,122 += +SEQUENCE_ID=pcr_primers_and_io +PRIMER_TASK=pick_pcr_primers_and_hyb_PROBE +SEQUENCE_TEMPLATE=NATCCANGCTGTTTCACACTTACGGAAATGGANGACTNCCCATAGCTNCTGGGCTNTTATCCAAAAAACATTCTCTNNNTGNTGCTTATTGGCNTCTCNAAGTGTGCNNACTTCATGGGCCTACACANAATTGNNCANGGGCTTAGCGTGCGTGGGGTCCCTNGGTAACATNGGCATCTCTGCACTTCNNGGTNNCCTTCCTTCTNACCAAGGGGTATCTGGNTGCCAGTGGCTTTTCTCCAACAGGAANGGGGGTATGTGCATTNGCAGACATCAATGANCATGAGAGATATNAATAGG += +SEQUENCE_ID=left_only +PRIMER_TASK=pick_left_only +SEQUENCE_TARGET=100,20 +SEQUENCE_TEMPLATE=GNATCTGCACCCTGTAAACCCTGGCTCCTGCCTCTCCGGGACACCCCACTGAGGTCAGCACACCCTGCAGGTTTAGAAGGGGTGTCTGGGTGCATTTGGTGACACCGCAGGCAGAGGGGGGACNCCACAGCCAGCTCTGCCCGCGGATGCCACGTCCATTTGCTTCAGCAGGATC += +SEQUENCE_ID=right_only +PRIMER_TASK=PICK_right_only +SEQUENCE_TARGET=90,10 +SEQUENCE_TEMPLATE=GANCAATGCANGTAATGANGTTGGCANAATTATGAATCGCAATCAAATTTTTTCTATGACCTGATAGAAACGCCAACTCACTGTATGAGTCTGTTGTCAAAGTCTAACTTCCAAAACGCAAAGCACGCTCCCGATC += +SEQUENCE_ID=hyb_probe_only +PRIMER_TASK=pick_HYB_probe_ONly +SEQUENCE_INTERNAL_EXCLUDED_REGION=90,20 +SEQUENCE_TEMPLATE=GCATCTGCACCCTGTAAACCCTGGCTCCTGCCTCTCCGGGACACCCCACTGAGGTCAGCACACCCTGCAGGTTTAGAAGGGGTGTCTGGGTGCATTTGGTGACACCGCAGGCAGAGGGGGGACNCCACAGCCAGCTCTGCCCGCGGATGCCACGTCCATTTGCTTCAGCAGGATC += +SEQUENCE_ID=left_only +PRIMER_MIN_SIZE=20 +PRIMER_MAX_SIZE=20 +PRIMER_TASK=pick_left_only +SEQUENCE_TEMPLATE=TTAGAAGGGGTGTCTGGGTG += +SEQUENCE_ID=right_only +PRIMER_TASK=pick_left_only +SEQUENCE_TEMPLATE=TTAGAAGGGGTGTCTGGGTG += +SEQUENCE_ID=hyb_probe_only +PRIMER_INTERNAL_MIN_SIZE=20 +PRIMER_INTERNAL_MAX_SIZE=20 +PRIMER_TASK=pick_left_only +SEQUENCE_TEMPLATE=TTAGAAGGGGTGTCTGGGTG += diff --git a/test/primer_windows_newlines_output b/test/primer_windows_newlines_output new file mode 100644 index 00000000..04b159df --- /dev/null +++ b/test/primer_windows_newlines_output @@ -0,0 +1,412 @@ +PRIMER_THERMODYNAMIC_OLIGO_ALIGNMENT=1 +PRIMER_TM_FORMULA=1 +PRIMER_SALT_CORRECTIONS=1 +PRIMER_SALT_DIVALENT=1.5 +PRIMER_MAX_SELF_ANY=12 +PRIMER_PAIR_MAX_COMPL_ANY=12 +PRIMER_MAX_POLY_X=100 +PRIMER_MAX_SELF_END=8 +PRIMER_PAIR_MAX_COMPL_END=8 +P3_FILE_FLAG=1 +PRIMER_EXPLAIN_FLAG=1 +PRIMER_TASK=pick_pcr_primers +PRIMER_DNTP_CONC=0.0 +SEQUENCE_ID=pcr_primers +SEQUENCE_TEMPLATE=TCCTACAGCTGTGGGAAAATGatgcgtacgatccatgctagctagctactatcgattagctatcatctactatcatctactatctactacacatctactatcatctacacacacacacacactaggccgctatatcatctagcGCCGCCATGTACACACTG +SEQUENCE_TARGET=21,122 +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_LEFT_EXPLAIN=considered 10, low tm 7, high hairpin stability 3, ok 0 +PRIMER_RIGHT_EXPLAIN=considered 1, ok 1 +PRIMER_PAIR_EXPLAIN=considered 0, ok 0 +PRIMER_LEFT_NUM_RETURNED=0 +PRIMER_RIGHT_NUM_RETURNED=0 +PRIMER_INTERNAL_NUM_RETURNED=0 +PRIMER_PAIR_NUM_RETURNED=0 += +SEQUENCE_ID=pcr_primers_and_io +PRIMER_TASK=pick_pcr_primers_and_hyb_PROBE +SEQUENCE_TEMPLATE=NATCCANGCTGTTTCACACTTACGGAAATGGANGACTNCCCATAGCTNCTGGGCTNTTATCCAAAAAACATTCTCTNNNTGNTGCTTATTGGCNTCTCNAAGTGTGCNNACTTCATGGGCCTACACANAATTGNNCANGGGCTTAGCGTGCGTGGGGTCCCTNGGTAACATNGGCATCTCTGCACTTCNNGGTNNCCTTCCTTCTNACCAAGGGGTATCTGGNTGCCAGTGGCTTTTCTCCAACAGGAANGGGGGTATGTGCATTNGCAGACATCAATGANCATGAGAGATATNAATAGG +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_LEFT_EXPLAIN=considered 281, too many Ns 210, low tm 32, high tm 27, ok 12 +PRIMER_RIGHT_EXPLAIN=considered 284, too many Ns 210, low tm 11, high tm 38, high hairpin stability 8, ok 17 +PRIMER_INTERNAL_EXPLAIN=considered 393, too many Ns 280, low tm 76, high tm 6, long poly-x seq 3, ok 28 +PRIMER_PAIR_EXPLAIN=considered 5, 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=3.564009 +PRIMER_LEFT_0_PENALTY=2.796318 +PRIMER_RIGHT_0_PENALTY=0.767691 +PRIMER_INTERNAL_0_PENALTY=0.180037 +PRIMER_LEFT_0_SEQUENCE=GCTGTTTCACACTTACGGAA +PRIMER_RIGHT_0_SEQUENCE=TGTTGGAGAAAAGCCACTGG +PRIMER_INTERNAL_0_SEQUENCE=TTAGCGTGCGTGGGGTCCCT +PRIMER_LEFT_0=7,20 +PRIMER_RIGHT_0=244,20 +PRIMER_INTERNAL_0=142,20 +PRIMER_LEFT_0_TM=57.204 +PRIMER_RIGHT_0_TM=59.232 +PRIMER_INTERNAL_0_TM=60.180 +PRIMER_LEFT_0_GC_PERCENT=45.000 +PRIMER_RIGHT_0_GC_PERCENT=50.000 +PRIMER_INTERNAL_0_GC_PERCENT=65.000 +PRIMER_INTERNAL_0_SELF_ANY_TH=5.01 +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=35.22 +PRIMER_INTERNAL_0_HAIRPIN_TH=34.96 +PRIMER_LEFT_0_END_STABILITY=4.3000 +PRIMER_RIGHT_0_END_STABILITY=4.0000 +PRIMER_PAIR_0_COMPL_ANY_TH=0.00 +PRIMER_PAIR_0_COMPL_END_TH=0.00 +PRIMER_PAIR_0_PRODUCT_SIZE=238 +PRIMER_PAIR_1_PENALTY=3.818870 +PRIMER_LEFT_1_PENALTY=3.051179 +PRIMER_RIGHT_1_PENALTY=0.767691 +PRIMER_INTERNAL_1_PENALTY=0.180037 +PRIMER_LEFT_1_SEQUENCE=GCTGTTTCACACTTACGGAAA +PRIMER_RIGHT_1_SEQUENCE=TGTTGGAGAAAAGCCACTGG +PRIMER_INTERNAL_1_SEQUENCE=TTAGCGTGCGTGGGGTCCCT +PRIMER_LEFT_1=7,21 +PRIMER_RIGHT_1=244,20 +PRIMER_INTERNAL_1=142,20 +PRIMER_LEFT_1_TM=57.949 +PRIMER_RIGHT_1_TM=59.232 +PRIMER_INTERNAL_1_TM=60.180 +PRIMER_LEFT_1_GC_PERCENT=42.857 +PRIMER_RIGHT_1_GC_PERCENT=50.000 +PRIMER_INTERNAL_1_GC_PERCENT=65.000 +PRIMER_INTERNAL_1_SELF_ANY_TH=5.01 +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=35.22 +PRIMER_INTERNAL_1_HAIRPIN_TH=34.96 +PRIMER_LEFT_1_END_STABILITY=3.1300 +PRIMER_RIGHT_1_END_STABILITY=4.0000 +PRIMER_PAIR_1_COMPL_ANY_TH=0.00 +PRIMER_PAIR_1_COMPL_END_TH=0.00 +PRIMER_PAIR_1_PRODUCT_SIZE=238 +PRIMER_PAIR_2_PENALTY=3.853878 +PRIMER_LEFT_2_PENALTY=2.796318 +PRIMER_RIGHT_2_PENALTY=1.057560 +PRIMER_INTERNAL_2_PENALTY=0.180037 +PRIMER_LEFT_2_SEQUENCE=GCTGTTTCACACTTACGGAA +PRIMER_RIGHT_2_SEQUENCE=TCCTGTTGGAGAAAAGCCAC +PRIMER_INTERNAL_2_SEQUENCE=TTAGCGTGCGTGGGGTCCCT +PRIMER_LEFT_2=7,20 +PRIMER_RIGHT_2=247,20 +PRIMER_INTERNAL_2=142,20 +PRIMER_LEFT_2_TM=57.204 +PRIMER_RIGHT_2_TM=58.942 +PRIMER_INTERNAL_2_TM=60.180 +PRIMER_LEFT_2_GC_PERCENT=45.000 +PRIMER_RIGHT_2_GC_PERCENT=50.000 +PRIMER_INTERNAL_2_GC_PERCENT=65.000 +PRIMER_INTERNAL_2_SELF_ANY_TH=5.01 +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=38.11 +PRIMER_INTERNAL_2_HAIRPIN_TH=34.96 +PRIMER_LEFT_2_END_STABILITY=4.3000 +PRIMER_RIGHT_2_END_STABILITY=5.0100 +PRIMER_PAIR_2_COMPL_ANY_TH=0.00 +PRIMER_PAIR_2_COMPL_END_TH=0.00 +PRIMER_PAIR_2_PRODUCT_SIZE=241 +PRIMER_PAIR_3_PENALTY=3.857112 +PRIMER_LEFT_3_PENALTY=2.796318 +PRIMER_RIGHT_3_PENALTY=1.060794 +PRIMER_INTERNAL_3_PENALTY=0.180037 +PRIMER_LEFT_3_SEQUENCE=GCTGTTTCACACTTACGGAA +PRIMER_RIGHT_3_SEQUENCE=CCTGTTGGAGAAAAGCCACT +PRIMER_INTERNAL_3_SEQUENCE=TTAGCGTGCGTGGGGTCCCT +PRIMER_LEFT_3=7,20 +PRIMER_RIGHT_3=246,20 +PRIMER_INTERNAL_3=142,20 +PRIMER_LEFT_3_TM=57.204 +PRIMER_RIGHT_3_TM=58.939 +PRIMER_INTERNAL_3_TM=60.180 +PRIMER_LEFT_3_GC_PERCENT=45.000 +PRIMER_RIGHT_3_GC_PERCENT=50.000 +PRIMER_INTERNAL_3_GC_PERCENT=65.000 +PRIMER_INTERNAL_3_SELF_ANY_TH=5.01 +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=35.22 +PRIMER_INTERNAL_3_HAIRPIN_TH=34.96 +PRIMER_LEFT_3_END_STABILITY=4.3000 +PRIMER_RIGHT_3_END_STABILITY=4.0000 +PRIMER_PAIR_3_COMPL_ANY_TH=0.00 +PRIMER_PAIR_3_COMPL_END_TH=0.00 +PRIMER_PAIR_3_PRODUCT_SIZE=240 +PRIMER_PAIR_4_PENALTY=4.080246 +PRIMER_LEFT_4_PENALTY=3.312555 +PRIMER_RIGHT_4_PENALTY=0.767691 +PRIMER_INTERNAL_4_PENALTY=0.180037 +PRIMER_LEFT_4_SEQUENCE=GCTGTTTCACACTTACGGAAATG +PRIMER_RIGHT_4_SEQUENCE=TGTTGGAGAAAAGCCACTGG +PRIMER_INTERNAL_4_SEQUENCE=TTAGCGTGCGTGGGGTCCCT +PRIMER_LEFT_4=7,23 +PRIMER_RIGHT_4=244,20 +PRIMER_INTERNAL_4=142,20 +PRIMER_LEFT_4_TM=59.687 +PRIMER_RIGHT_4_TM=59.232 +PRIMER_INTERNAL_4_TM=60.180 +PRIMER_LEFT_4_GC_PERCENT=43.478 +PRIMER_RIGHT_4_GC_PERCENT=50.000 +PRIMER_INTERNAL_4_GC_PERCENT=65.000 +PRIMER_INTERNAL_4_SELF_ANY_TH=5.01 +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=34.24 +PRIMER_RIGHT_4_HAIRPIN_TH=35.22 +PRIMER_INTERNAL_4_HAIRPIN_TH=34.96 +PRIMER_LEFT_4_END_STABILITY=2.3200 +PRIMER_RIGHT_4_END_STABILITY=4.0000 +PRIMER_PAIR_4_COMPL_ANY_TH=0.00 +PRIMER_PAIR_4_COMPL_END_TH=0.00 +PRIMER_PAIR_4_PRODUCT_SIZE=238 += +SEQUENCE_ID=left_only +PRIMER_TASK=pick_left_only +SEQUENCE_TARGET=100,20 +SEQUENCE_TEMPLATE=GNATCTGCACCCTGTAAACCCTGGCTCCTGCCTCTCCGGGACACCCCACTGAGGTCAGCACACCCTGCAGGTTTAGAAGGGGTGTCTGGGTGCATTTGGTGACACCGCAGGCAGAGGGGGGACNCCACAGCCAGCTCTGCCCGCGGATGCCACGTCCATTTGCTTCAGCAGGATC +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_LEFT_EXPLAIN=considered 1121, too many Ns 38, overlap target 23, low tm 34, high tm 809, high hairpin stability 88, ok 129 +PRIMER_LEFT_NUM_RETURNED=5 +PRIMER_RIGHT_NUM_RETURNED=0 +PRIMER_INTERNAL_NUM_RETURNED=0 +PRIMER_PAIR_NUM_RETURNED=0 +PRIMER_LEFT_0_PENALTY=0.199248 +PRIMER_LEFT_0_SEQUENCE=TGCAGGTTTAGAAGGGGTGT +PRIMER_LEFT_0=65,20 +PRIMER_LEFT_0_TM=59.801 +PRIMER_LEFT_0_GC_PERCENT=50.000 +PRIMER_LEFT_0_SELF_ANY_TH=0.00 +PRIMER_LEFT_0_SELF_END_TH=0.00 +PRIMER_LEFT_0_HAIRPIN_TH=0.00 +PRIMER_LEFT_0_END_STABILITY=4.1600 +PRIMER_LEFT_1_PENALTY=0.243148 +PRIMER_LEFT_1_SEQUENCE=CCTGCAGGTTTAGAAGGGGT +PRIMER_LEFT_1=63,20 +PRIMER_LEFT_1_TM=60.243 +PRIMER_LEFT_1_GC_PERCENT=55.000 +PRIMER_LEFT_1_SELF_ANY_TH=26.16 +PRIMER_LEFT_1_SELF_END_TH=0.00 +PRIMER_LEFT_1_HAIRPIN_TH=0.00 +PRIMER_LEFT_1_END_STABILITY=4.9500 +PRIMER_LEFT_2_PENALTY=0.382831 +PRIMER_LEFT_2_SEQUENCE=ATGCCACGTCCATTTGCTTC +PRIMER_LEFT_2=146,20 +PRIMER_LEFT_2_TM=60.383 +PRIMER_LEFT_2_GC_PERCENT=50.000 +PRIMER_LEFT_2_SELF_ANY_TH=0.00 +PRIMER_LEFT_2_SELF_END_TH=0.00 +PRIMER_LEFT_2_HAIRPIN_TH=0.00 +PRIMER_LEFT_2_END_STABILITY=3.8600 +PRIMER_LEFT_3_PENALTY=0.382831 +PRIMER_LEFT_3_SEQUENCE=GATGCCACGTCCATTTGCTT +PRIMER_LEFT_3=145,20 +PRIMER_LEFT_3_TM=60.383 +PRIMER_LEFT_3_GC_PERCENT=50.000 +PRIMER_LEFT_3_SELF_ANY_TH=0.92 +PRIMER_LEFT_3_SELF_END_TH=0.00 +PRIMER_LEFT_3_HAIRPIN_TH=0.00 +PRIMER_LEFT_3_END_STABILITY=3.9100 +PRIMER_LEFT_4_PENALTY=0.417433 +PRIMER_LEFT_4_SEQUENCE=TTAGAAGGGGTGTCTGGGTG +PRIMER_LEFT_4=72,20 +PRIMER_LEFT_4_TM=59.583 +PRIMER_LEFT_4_GC_PERCENT=55.000 +PRIMER_LEFT_4_SELF_ANY_TH=0.00 +PRIMER_LEFT_4_SELF_END_TH=0.00 +PRIMER_LEFT_4_HAIRPIN_TH=0.00 +PRIMER_LEFT_4_END_STABILITY=4.6100 += +SEQUENCE_ID=right_only +PRIMER_TASK=PICK_right_only +SEQUENCE_TARGET=90,10 +SEQUENCE_TEMPLATE=GANCAATGCANGTAATGANGTTGGCANAATTATGAATCGCAATCAAATTTTTTCTATGACCTGATAGAAACGCCAACTCACTGTATGAGTCTGTTGTCAAAGTCTAACTTCCAAAACGCAAAGCACGCTCCCGATC +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_RIGHT_EXPLAIN=considered 623, too many Ns 27, overlap target 36, GC content failed 5, low tm 279, high tm 102, high hairpin stability 25, ok 149 +PRIMER_LEFT_NUM_RETURNED=0 +PRIMER_RIGHT_NUM_RETURNED=5 +PRIMER_INTERNAL_NUM_RETURNED=0 +PRIMER_PAIR_NUM_RETURNED=0 +PRIMER_RIGHT_0_PENALTY=0.033659 +PRIMER_RIGHT_0_SEQUENCE=CGTGCTTTGCGTTTTGGAAG +PRIMER_RIGHT_0=126,20 +PRIMER_RIGHT_0_TM=60.034 +PRIMER_RIGHT_0_GC_PERCENT=50.000 +PRIMER_RIGHT_0_SELF_ANY_TH=0.00 +PRIMER_RIGHT_0_SELF_END_TH=0.00 +PRIMER_RIGHT_0_HAIRPIN_TH=0.00 +PRIMER_RIGHT_0_END_STABILITY=3.4600 +PRIMER_RIGHT_1_PENALTY=1.023063 +PRIMER_RIGHT_1_SEQUENCE=GTGCTTTGCGTTTTGGAAGT +PRIMER_RIGHT_1=125,20 +PRIMER_RIGHT_1_TM=58.977 +PRIMER_RIGHT_1_GC_PERCENT=45.000 +PRIMER_RIGHT_1_SELF_ANY_TH=0.00 +PRIMER_RIGHT_1_SELF_END_TH=0.00 +PRIMER_RIGHT_1_HAIRPIN_TH=0.00 +PRIMER_RIGHT_1_END_STABILITY=3.0100 +PRIMER_RIGHT_2_PENALTY=1.026309 +PRIMER_RIGHT_2_SEQUENCE=GAGCGTGCTTTGCGTTTTG +PRIMER_RIGHT_2=129,19 +PRIMER_RIGHT_2_TM=60.026 +PRIMER_RIGHT_2_GC_PERCENT=52.632 +PRIMER_RIGHT_2_SELF_ANY_TH=0.00 +PRIMER_RIGHT_2_SELF_END_TH=0.00 +PRIMER_RIGHT_2_HAIRPIN_TH=37.77 +PRIMER_RIGHT_2_END_STABILITY=2.4400 +PRIMER_RIGHT_3_PENALTY=1.363507 +PRIMER_RIGHT_3_SEQUENCE=GTGCTTTGCGTTTTGGAAGTT +PRIMER_RIGHT_3=125,21 +PRIMER_RIGHT_3_TM=59.636 +PRIMER_RIGHT_3_GC_PERCENT=42.857 +PRIMER_RIGHT_3_SELF_ANY_TH=0.00 +PRIMER_RIGHT_3_SELF_END_TH=0.00 +PRIMER_RIGHT_3_HAIRPIN_TH=0.00 +PRIMER_RIGHT_3_END_STABILITY=2.6600 +PRIMER_RIGHT_4_PENALTY=1.754055 +PRIMER_RIGHT_4_SEQUENCE=TGCTTTGCGTTTTGGAAGTT +PRIMER_RIGHT_4=124,20 +PRIMER_RIGHT_4_TM=58.246 +PRIMER_RIGHT_4_GC_PERCENT=40.000 +PRIMER_RIGHT_4_SELF_ANY_TH=0.00 +PRIMER_RIGHT_4_SELF_END_TH=0.00 +PRIMER_RIGHT_4_HAIRPIN_TH=0.00 +PRIMER_RIGHT_4_END_STABILITY=2.6600 += +SEQUENCE_ID=hyb_probe_only +PRIMER_TASK=pick_HYB_probe_ONly +SEQUENCE_INTERNAL_EXCLUDED_REGION=90,20 +SEQUENCE_TEMPLATE=GCATCTGCACCCTGTAAACCCTGGCTCCTGCCTCTCCGGGACACCCCACTGAGGTCAGCACACCCTGCAGGTTTAGAAGGGGTGTCTGGGTGCATTTGGTGACACCGCAGGCAGAGGGGGGACNCCACAGCCAGCTCTGCCCGCGGATGCCACGTCCATTTGCTTCAGCAGGATC +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_INTERNAL_EXPLAIN=considered 1040, too many Ns 27, overlap excluded region 33, low tm 271, high tm 297, high hairpin stability 144, ok 268 +PRIMER_LEFT_NUM_RETURNED=0 +PRIMER_RIGHT_NUM_RETURNED=0 +PRIMER_INTERNAL_NUM_RETURNED=5 +PRIMER_PAIR_NUM_RETURNED=0 +PRIMER_INTERNAL_0_PENALTY=0.031870 +PRIMER_INTERNAL_0_SEQUENCE=GCTCCTGCCTCTCCGGGACA +PRIMER_INTERNAL_0=23,20 +PRIMER_INTERNAL_0_TM=59.968 +PRIMER_INTERNAL_0_GC_PERCENT=70.000 +PRIMER_INTERNAL_0_SELF_ANY_TH=0.00 +PRIMER_INTERNAL_0_SELF_END_TH=0.00 +PRIMER_INTERNAL_0_HAIRPIN_TH=30.41 +PRIMER_INTERNAL_1_PENALTY=0.111561 +PRIMER_INTERNAL_1_SEQUENCE=CCGCGGATGCCACGTCCATT +PRIMER_INTERNAL_1=140,20 +PRIMER_INTERNAL_1_TM=60.112 +PRIMER_INTERNAL_1_GC_PERCENT=65.000 +PRIMER_INTERNAL_1_SELF_ANY_TH=20.33 +PRIMER_INTERNAL_1_SELF_END_TH=0.00 +PRIMER_INTERNAL_1_HAIRPIN_TH=35.40 +PRIMER_INTERNAL_2_PENALTY=0.388807 +PRIMER_INTERNAL_2_SEQUENCE=TCCTGCCTCTCCGGGACACC +PRIMER_INTERNAL_2=25,20 +PRIMER_INTERNAL_2_TM=59.611 +PRIMER_INTERNAL_2_GC_PERCENT=70.000 +PRIMER_INTERNAL_2_SELF_ANY_TH=0.00 +PRIMER_INTERNAL_2_SELF_END_TH=0.00 +PRIMER_INTERNAL_2_HAIRPIN_TH=30.41 +PRIMER_INTERNAL_3_PENALTY=0.602519 +PRIMER_INTERNAL_3_SEQUENCE=CCTGCCTCTCCGGGACACCC +PRIMER_INTERNAL_3=26,20 +PRIMER_INTERNAL_3_TM=60.603 +PRIMER_INTERNAL_3_GC_PERCENT=75.000 +PRIMER_INTERNAL_3_SELF_ANY_TH=0.00 +PRIMER_INTERNAL_3_SELF_END_TH=0.00 +PRIMER_INTERNAL_3_HAIRPIN_TH=38.95 +PRIMER_INTERNAL_4_PENALTY=1.037103 +PRIMER_INTERNAL_4_SEQUENCE=CCACAGCCAGCTCTGCCCG +PRIMER_INTERNAL_4=124,19 +PRIMER_INTERNAL_4_TM=60.037 +PRIMER_INTERNAL_4_GC_PERCENT=73.684 +PRIMER_INTERNAL_4_SELF_ANY_TH=0.00 +PRIMER_INTERNAL_4_SELF_END_TH=0.00 +PRIMER_INTERNAL_4_HAIRPIN_TH=34.17 += +SEQUENCE_ID=left_only +PRIMER_MIN_SIZE=20 +PRIMER_MAX_SIZE=20 +PRIMER_TASK=pick_left_only +SEQUENCE_TEMPLATE=TTAGAAGGGGTGTCTGGGTG +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_LEFT_EXPLAIN=considered 1, ok 1 +PRIMER_LEFT_NUM_RETURNED=1 +PRIMER_RIGHT_NUM_RETURNED=0 +PRIMER_INTERNAL_NUM_RETURNED=0 +PRIMER_PAIR_NUM_RETURNED=0 +PRIMER_LEFT_0_PENALTY=0.417433 +PRIMER_LEFT_0_SEQUENCE=TTAGAAGGGGTGTCTGGGTG +PRIMER_LEFT_0=0,20 +PRIMER_LEFT_0_TM=59.583 +PRIMER_LEFT_0_GC_PERCENT=55.000 +PRIMER_LEFT_0_SELF_ANY_TH=0.00 +PRIMER_LEFT_0_SELF_END_TH=0.00 +PRIMER_LEFT_0_HAIRPIN_TH=0.00 +PRIMER_LEFT_0_END_STABILITY=4.6100 += +SEQUENCE_ID=right_only +PRIMER_TASK=pick_left_only +SEQUENCE_TEMPLATE=TTAGAAGGGGTGTCTGGGTG +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_LEFT_EXPLAIN=considered 1, ok 1 +PRIMER_LEFT_NUM_RETURNED=1 +PRIMER_RIGHT_NUM_RETURNED=0 +PRIMER_INTERNAL_NUM_RETURNED=0 +PRIMER_PAIR_NUM_RETURNED=0 +PRIMER_LEFT_0_PENALTY=0.417433 +PRIMER_LEFT_0_SEQUENCE=TTAGAAGGGGTGTCTGGGTG +PRIMER_LEFT_0=0,20 +PRIMER_LEFT_0_TM=59.583 +PRIMER_LEFT_0_GC_PERCENT=55.000 +PRIMER_LEFT_0_SELF_ANY_TH=0.00 +PRIMER_LEFT_0_SELF_END_TH=0.00 +PRIMER_LEFT_0_HAIRPIN_TH=0.00 +PRIMER_LEFT_0_END_STABILITY=4.6100 += +SEQUENCE_ID=hyb_probe_only +PRIMER_INTERNAL_MIN_SIZE=20 +PRIMER_INTERNAL_MAX_SIZE=20 +PRIMER_TASK=pick_left_only +SEQUENCE_TEMPLATE=TTAGAAGGGGTGTCTGGGTG +PRIMER_WARNING=PRIMER_SALT_DIVALENT > 0.0 but PRIMER_DNTP_CONC <= 0.0; use reasonable value for PRIMER_DNTP_CONC +PRIMER_LEFT_EXPLAIN=considered 1, ok 1 +PRIMER_LEFT_NUM_RETURNED=1 +PRIMER_RIGHT_NUM_RETURNED=0 +PRIMER_INTERNAL_NUM_RETURNED=0 +PRIMER_PAIR_NUM_RETURNED=0 +PRIMER_LEFT_0_PENALTY=0.417433 +PRIMER_LEFT_0_SEQUENCE=TTAGAAGGGGTGTCTGGGTG +PRIMER_LEFT_0=0,20 +PRIMER_LEFT_0_TM=59.583 +PRIMER_LEFT_0_GC_PERCENT=55.000 +PRIMER_LEFT_0_SELF_ANY_TH=0.00 +PRIMER_LEFT_0_SELF_END_TH=0.00 +PRIMER_LEFT_0_HAIRPIN_TH=0.00 +PRIMER_LEFT_0_END_STABILITY=4.6100 +=