update

primer3.nf

@@ -10,12 +10,12 @@ process primer3_index {
     output:
       path "${fasta}*.list"
     shell:
-      '''
+'''
 glistmaker !{fasta} -w 11          
 glistmaker !{fasta} -w 16          
 ln out_11.list !{fasta}_11.list
 ln out_16.list !{fasta}_16.list
-      '''
+'''
 }
 
 process primer3_conf {
@@ -29,21 +29,28 @@ process primer3_conf {
       path "custom_primer3.conf"
     shell:
 '''
+##The MASK_KMERLIST does give an error if run with a dummy value, so expect it is working?
+## See the Primer3 manual for details on these parameters
+
+##The "SEQUENCE_TARGET" parameter is fairly important, as that ensures the PCR product goes across the gRNA site
+## The SEQUENCE_EXCLUDED_REGION ensures it doesn't pick a product right on the gRNA site
 
 echo "SEQUENCE_ID=$(seqkit fx2tab !{target} | cut -f 1)
 SEQUENCE_TEMPLATE=$(seqkit fx2tab !{target} | cut -f 2)
+SEQUENCE_TARGET=500,20
+SEQUENCE_EXCLUDED_REGION=465,70
 PRIMER_TASK=generic
 PRIMER_PICK_LEFT_PRIMER=1
 PRIMER_PICK_INTERNAL_OLIGO=0
 PRIMER_PICK_RIGHT_PRIMER=1
 PRIMER_OPT_SIZE=20
 PRIMER_MIN_SIZE=18
 PRIMER_MAX_SIZE=22
-PRIMER_PRODUCT_SIZE_RANGE=75-150
+PRIMER_PRODUCT_SIZE_RANGE=75-500
 PRIMER_EXPLAIN_FLAG=1
 PRIMER_MASK_TEMPLATE=1
-PRIMER_MASK_KMERLIST_PATH=./kmer_lists/
 PRIMER_MASK_KMERLIST_PREFIX=!{fasta}
+PRIMER_MASK_KMERLIST_PATH=./kmer_lists/
 =" > custom_primer3.conf      
 '''
 }
@@ -57,16 +64,44 @@ process primer3_calc {
       path conf
       path kmer_lists    
     output:
-      tuple path("${fasta}.crispr.db"),path("${fasta}.crispr.db.header")
+      path "primer3_results.txt"
     shell:
 '''
 mkdir -p kmer_lists
 ln ./*.list kmer_lists
+
+ID=$(head -n 1 !{conf} | cut -f 2 -d '=')
+
 primer3_core !{conf} > primer3_results.txt
 '''
 }
 
+process primer3_results2fasta {
+input:
+ path results
+shell:
+'''
+#!/usr/bin/env python
+import re
+import os
+import os.path
+print("hello world!")
 
+with open("!{results}", "r") as input_handle:
+    data = input_handle.read()
+##print(data)
+
+id_regex = 'SEQUENCE_ID=(.+)'
+bespoke_regex='(?P<id>PRIMER_PAIR_[0-9]+)_PENALTY=(?P<penalty>[0-9.]+).+?PRIMER_LEFT_[0-9]+_SEQUENCE=(?P<left>[atcgATCGnN]+).+?PRIMER_RIGHT_[0-9]+_SEQUENCE=(?P<right>[atcgATCGnN]+).+?PRIMER_PAIR_[0-9]+_PRODUCT_TM=[0-9.]+'
+
+matches = list(re.finditer(bespoke_regex,data,flags=re.MULTILINE|re.DOTALL))
+
+output_handle = open("!{results}.fa", "w")
+for m in matches:
+    record_id = ">
+
+'''
+}
 
 workflow {
   ref = channel.fromPath(params.genome)
@@ -75,4 +110,6 @@ workflow {
   primer3_conf(ref,target)
   primer3_index(ref)
   primer3_calc(primer3_conf.out,primer3_index.out)
+  primer3_results2fasta(primer3_calc.out)
+
 }