awk? how to retrieve info from massive Primer3 output file

Question

[sp]

Hi All,

I would like to make primer file in fasta format retrieving some information (Primer ID, and left & right sequence) from primer3 output as below. What would be the most efficient way to do this job? I figured that "AWK" might be a good tool for this, but no clue where to start. Mucho appreciate if any expert can help me!!!

from primer3 out:

{'PRIMER_INTERNAL_EXPLAIN': 'considered 3622, unacceptable product size 3492, ok 130',
'PRIMER_INTERNAL_NUM_RETURNED': 0L,
'PRIMER_LEFT_0': (124L, 19L),
'PRIMER_LEFT_0_END_STABILITY': 3.58,
'PRIMER_LEFT_0_GC_PERCENT': 52.63157894736842,
'PRIMER_LEFT_0_HAIRPIN_TH': 44.35240092833908,
'PRIMER_LEFT_0_PENALTY': 80.38508071355189,
'PRIMER_LEFT_0_SELF_ANY_TH': 0.0,
'PRIMER_LEFT_0_SELF_END_TH': 0.0,
'PRIMER_LEFT_0_SEQUENCE': '**CCTCAAGGTCTTCCTGTCA**',
'PRIMER_LEFT_0_TM': 55.953805840088194,
'PRIMER_LEFT_EXPLAIN': 'considered 598, overlap excluded region 64, GC content failed 278, low tm 14, high tm 155, high hairpin stability 33, long poly-x seq 16, ok 38',
'PRIMER_LEFT_NUM_RETURNED': 1L,
'PRIMER_PAIR_0_COMPL_ANY_TH': 0.0,
'PRIMER_PAIR_0_COMPL_END_TH': 0.0,
'PRIMER_PAIR_0_PENALTY': 789.1692592626196,
'PRIMER_PAIR_0_PRODUCT_SIZE': 76L,
'PRIMER_PAIR_NUM_RETURNED': 1L,
'PRIMER_RIGHT_0': (199L, 18L),
'PRIMER_RIGHT_0_END_STABILITY': 4.57,
'PRIMER_RIGHT_0_GC_PERCENT': 55.55555555555556,
'PRIMER_RIGHT_0_HAIRPIN_TH': 0.0,
'PRIMER_RIGHT_0_PENALTY': 108.78417854906772,
'PRIMER_RIGHT_0_SELF_ANY_TH': 10.083235050216501,
'PRIMER_RIGHT_0_SELF_END_TH': 0.0,
'PRIMER_RIGHT_0_SEQUENCE': '**GAAGTATACGCGGGCACA**',
'PRIMER_RIGHT_0_TM': 57.180918004325235,
'PRIMER_RIGHT_EXPLAIN': 'considered 658, overlap excluded region 63, GC content failed 314, low tm 14, high tm 179, high hairpin stability 20, long poly-x seq 3, ok 65',
'PRIMER_RIGHT_NUM_RETURNED': 1L,
'SEQUENCE_ID': '**chr10:43102245-43102445**',
'SEQUENCE_TEMPLATE': 'GGGTTTACACCAGCCCTGGAGCTCCTGCCTCCTCCCCATTCCCGACTGCCTGGCAGATGTGGCCGATGCCCCCACAGACCTGACTTCTCTCTGCAGACCGCGGCTTTCCCCTGCTCACCGTCTACCTCAAGGTCTTCCTGTCACCCACATCCCTTCGTGAGGGCGAGTGCCAGTGGCCAGGCTGTGCCCGCGTATACTTC'}
None

{'PRIMER_INTERNAL_EXPLAIN': 'considered 4710, unacceptable product size 2919, ok 1791',

'PRIMER_INTERNAL_NUM_RETURNED': 0L,
'PRIMER_LEFT_0': (52L, 18L),
'PRIMER_LEFT_0_END_STABILITY': 3.41,
'PRIMER_LEFT_0_GC_PERCENT': 61.111111111111114,
'PRIMER_LEFT_0_HAIRPIN_TH': 0.0,
'PRIMER_LEFT_0_PENALTY': 165.23988901926742,
'PRIMER_LEFT_0_SELF_ANY_TH': 0.0,
'PRIMER_LEFT_0_SELF_END_TH': 0.0,
'PRIMER_LEFT_0_SEQUENCE': '**CCATCTCGCCTGCACTGA**',
'PRIMER_LEFT_0_TM': 59.41918527210419,
'PRIMER_LEFT_EXPLAIN': 'considered 509, overlap excluded region 64, GC content failed 174, low tm 20, high tm 132, long poly-x seq 30, ok 89',
'PRIMER_LEFT_NUM_RETURNED': 1L,
'PRIMER_PAIR_0_COMPL_ANY_TH': 9.428746428806335,
'PRIMER_PAIR_0_COMPL_END_TH': 6.6828990446157945,
'PRIMER_PAIR_0_PENALTY': 367.4410290971364,
'PRIMER_PAIR_0_PRODUCT_SIZE': 96L,
'PRIMER_PAIR_NUM_RETURNED': 1L,
'PRIMER_RIGHT_0': (147L, 20L),
'PRIMER_RIGHT_0_END_STABILITY': 4.18,
'PRIMER_RIGHT_0_GC_PERCENT': 55.0,
'PRIMER_RIGHT_0_HAIRPIN_TH': 0.0,
'PRIMER_RIGHT_0_PENALTY': 102.20114007786896,
'PRIMER_RIGHT_0_SELF_ANY_TH': 21.29971050510221,
'PRIMER_RIGHT_0_SELF_END_TH': 3.95513181391766,
'PRIMER_RIGHT_0_SEQUENCE': '**CTGATGCAGGTACCACGTCT**',
'PRIMER_RIGHT_0_TM': 59.467426718579304,
'PRIMER_RIGHT_EXPLAIN': 'considered 562, overlap excluded region 64, GC content failed 239, high tm 175, high hairpin stability 12, long poly-x seq 24, ok 48',
'PRIMER_RIGHT_NUM_RETURNED': 1L,
'SEQUENCE_ID': '**chr10:43106282-43106482**',
'SEQUENCE_TEMPLATE': 'GTGTGGGACGTGCAGCATTCTAAGGTCTCTGGTTTTGGGGGGTCTGAGGGGCCCATCTCGCCTGCACTGACCAACGCCCTCTGCATCCTGCAGGACACCGTGGTGGCCACGCTGCGTGTCTTCGATGCAGACGTGGTACCTGCATCAGGGGAGCTGGTGAGGCGGTACACAAGCACGCTGCTCCCCGGGGACACCTGGGC'}
None

and so on...

To desired primer format:

>chr10:43102245-43102445-left
CCTCAAGGTCTTCCTGTCA
>chr10:43102245-43102445-right
GAAGTATACGCGGGCACA
>chr10:43106282-43106482-left
CCATCTCGCCTGCACTGA
>chr10:43106282-43106482-right
CTGATGCAGGTACCACGTCT

etc...

[sp]

I ran your script just now, and encountered error as below.

sp@sp-VBox:~/shared$ grep -E "PRIMER_RIGHT_0_SEQUENCE|PRIMER_LEFT_0_SEQUENCE|SEQUENCE_ID" **chr10_Primers_Full.out** | paste - - - | awk '{ gsub("\047|,","",$0); print ">"$6"-left\n"$2"\n" ">"$6"-right\n"$4}'​ **> output.fa**

awk: cmd. line:1: { gsub("\047|,","",$0); print ">"$6"-left\n"$2"\n" ">"$6"-right\n"$4}​
awk: cmd. line:1:                                                                      ^ invalid char '�' in expression

Any suggestion?

[GouthamAtla]

using windows ?

[sp]

Copied your code from windows and pasted it virtual ubuntu machine.

Answer 1

[GouthamAtla]

grep -E "PRIMER_RIGHT_0_SEQUENCE|PRIMER_LEFT_0_SEQUENCE|SEQUENCE_ID" test.fasta |\
     paste - - - |\
     awk '{ gsub("\047|,","",$0); print `">"$6"-left\n"$2"\n"` ">"$6"-right\n"$4}'​

Assumptions:

1. The right,left sequences and sequence ID follows same order for every record/primer.
2. The left and right sequence is always present in a single line.

It could be more elegant.

[sp]

I typed your script on the terminal directly and it worked nicely.

Mucho gracias!

Answer 2

[Charles Plessy]

For a simpler output format, have a look at the eprimer32 wrapper in EMBOSS.