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readme.md

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@@ -58,16 +58,18 @@ optional arguments:
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```
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Use _C. nigoni_ and _C.briggsae_ genomes as example. The two fasta files can be downloaded separately
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from [cb4.fa](https://github.com/Runsheng/cbgenome/releases/download/cb5pre_cn3pre/cb5.fa.gz) and
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from [cb5.fa](https://github.com/Runsheng/cbgenome/releases/download/cb5pre_cn3pre/cb5.fa.gz) and
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[cn3_new.fa](https://github.com/Runsheng/cbgenome/releases/download/cb5pre_cn3pre/cn3_new.fa.gz).
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The _C. nigoni_ genome is cn3_new.fa and _C. briggsae_ genome is cb5.fa. To design _C. briggsae_ unique primer,
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which would not amplify any region in _C. nigoni_, and amplify only one region in _C. briggsae_.
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The targeted region for C. briggsae is ChrX:12881200-15106660 (-pos),
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one primer is designed for every 4kb interval (--interval).
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```
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```bash
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primerdesign.py -g1 cb5.fa -g2 cn3_new.fa -pos "ChrX:12881200-15106660" --interval 4000
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```
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```
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# check the result in file "primers_ChrX:12881200-15106660.txt"
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head primers_ChrX\:12881200-15106660.txt
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#ChrX:12881200-12881700 GATCCAAAACATGAGTGGCC CGAGATCATTGGCTCAAAGT 287
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```bash
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ispcr.py -f gcactttcatgtccctcaac -r cactctattctcaccccacc -g cb5.fa -o ispcr.fa
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```
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```
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# check the PCR product in ispcr.fa
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head ispcr.fa
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#>ChrI:230699-231076_RC
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#GCACTTTCATGTCCCTCAACCAGTCGTTTTTCCTTACCTCTCCCCTTCCTTTTTTCCCCCTCCCAGATGACGTCACCCATCTGTCC
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## Roadmap for other functions:
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1. To use user-provided primer parameters.Primer design parameter now is fine-tuned for general purpose PCR, which can be found in "general_settings.py".This file may need be modified to generate primers for specific purpose PCR like real-time qPCR.
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2. To update the RFLP method for primer design to differ sequences with almost identical sequence.
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3. To update the primer design using VCF file.
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3. To update the primer design using VCF file for closely related haplotypes.
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