HT-16srRNA.GABA

HT-16srRNA.GABA is a comprehensive bioinformatics pipeline designed for the high-throughput identification of microbial strains using 16S rRNA gene sequencing data. The pipeline includes multiple steps such as duplex sequencing, taxonomic identification, and abundance profiling.

1. Install dependencies

#create a environment with conda
conda create --name emu
#install emu seqkit blast 
conda install -c bioconda emu seqkit blast samtools
git clone https://github.com/DingDingFan/HT-16srRNA.GABA.git
cd HT-16srRNA.GABA

2. How to run

step 1: activate environment

conda activate emu

step 2: covert fastq to fasta formate

seqkit fq2fa pass.fq.gz  > pass.fq.fasta 
samtools faidx pass.fq.fasta

step 3: Reads aligend to barcode database

sh  script/blastn.single.sh pass.fq.fasta  script/50X50barcode.fasta aln # the database is custom

step 4: Duplex and add sample information

perl script/deal.pfa.pl pass.fq.fasta.fai   aln.blast  > aln.blast.deplex
perl script/add.sample.pl dual_barcode_pair.txt  aln.blast.deplex > aln.blast.demultiplex.add.sample.xls
 # "dual_barcode_pair.txt" is a file that contains a mapping of samples to their corresponding barcodes in a sheet format
for i in deplex/*/seq.id
    echo "head -1000 $i | perl ~/bin/commom_bin/fishInWinter.pl --ff fasta /dev/stdin pass.fq.fasta >$i.fas  "
end > batch.getseq.sh    
perl script/commom_bin/multi-process.pl  -cpu 20 batch.getseq.sh

step 5: taxonomic identification and generation of abundance profiles

for i in $PWD/deplex/*
do
echo "cd $i; sh script/nano.emu.sh "
done
end  >batch.sh
perl ~/bin/commom_bin/multi-process.pl  -cpu 10  batch.sh

step 6 Merge all result

for i in deplex/*/seq.id_rel-abundance.tsv
do
perl ~/bin/meta/got.emu.result.pl $i 
done > Nanopore.assign.xls

others

Nanopore sequencing test data can be download form Bioproject(PRJNA1170189). ohters Pacbio/HIFI NGS also can be downloaded . sanger sequencing attached in the demo dirment.

License

This project is licensed under the MIT License

Contact

For questions or inquiries, please contact [biocomfun@qq.com].