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Cyclone-DNAbarcode-COI

DESCRIPTION

A set of tootkit for dealing with COI amplicons using Cyclone sequencing platform

INSTALLATION

  • Clone from github
$ git clone https://github.com/BioEarthDigital/CycCOI.git

Requirements

(1) python modules

from Bio import SeqIO
from Bio.Seq import Seq
from Bio.SeqRecord import SeqRecord
import numpy as np
from collections import defaultdict
import argparse
import logging
import sys
import os
from multiprocessing import Pool
import multiprocessing
from icecream import ic

DATA requirements:

(1) CycloneSEQ reads

(2) primers list

primer.list

for	TAAACTTCTGGATGTCCAAAAAATCA
rev	TTTCAACAAATCATAAAGATATTGG

(3) index(barcodes for identifying samples in which plate) list

plate.index.tsv

P1	TCGGTCTTAGACG
P2	TGTGAAGTTGCCA
P3	AGATTCTACACAA
P4	ATGCGATTAATTG
P5	GGCTGTTACAACA

(4) index(barcodes for identifying samples in a plate) list

cell.index.tsv

001     AAAGC 
002     AACAG 
003     AACCT 
004     AACTC 
005     AAGCA

Get start

  1. QC, filtering sequencing by length, gc and generate report figures
python3 bin/CycFqFilter.py -q 7 -l 700 -L 770 -g 0.2 -G 0.6 -o test.clean test.fastq.gz
  1. assign sequencing by plate index and well index
$ python3 ../bin/pcr_demultiplex.py -p primer.txt --plate-index plate.index.tsv --well-index cell.index.tsv -f test.fa  -o out  --primer_max_mismatch 3 --primer_max_indel 3 --index_max_mismatch 2 --index_max_indel 1
  1. make consensus sequence for each well
python3 ../bin/cluster_consensus.py -i assigned.fa -o cluster --id_threshold 0.95 --keep_temp_files