A script for sgRNA extraction and quality control.
Clone this repo:
git clone https://github.com/Jianhua-Wang/LibraryQCset up conda environment and activate the environment:
cd LibraryQC
conda env create -f libraryqccondaenv.yml
conda activate libraryqc(base) ➜ LibraryQC git:(master) ✗ python libraryqc.py -h
usage: Librayqc using pattern matching to extract the sgRNA from pair-end sequencing data.
python libraryqc.py -i input/file_path.csv -o output -v input/vector.fa -l
input/library.csv
optional arguments:
-h, --help show this help message and exit
-i , --input csv file contains the sampl name and fq1, fq2 file path of
samples
-o , --output directory of output file
-v , --vector the fasta file of your vector, containing two sequences, 5'
and 3'.
-l , --library csv file of sgRNA you designed
-s , --shot the number of bases upstream and downstream of sgRNA you
exptected to match. More shot might reduce the false
positive. default=4The path of fastq files are specified in a csv file, for example:
| sample_name | fq_1 | fq_2 |
|---|---|---|
| day0 | ./input/data/day0_1.fq.gz | ./input/data/day0_1.fq.gz |
| day14 | ./input/data/day14_1.fq.gz | ./input/data/day14_1.fq.gz |
| vf | ./input/data/vf_1.fq.gz | ./input/data/vf_1.fq.gz |
The library you designed or download from other website, for example:
| id | sequence | note |
|---|---|---|
| 1 | ATAGGCACACATGAAGCGGA | |
| 2 | TTTGCTGATAACTAGATCTA | |
| 3 | TTGCAGGCCGCGATCTGTGC | |
| 4 | GGTGGAGACTCCGAGTGTAG | |
| 5 | ACTTCAAGTACGAGAACCAG | - |
| 6 | AATTGCATTCGGTTTCTATC | - |
| 7 | GAGCTTTTTGGGGTGTGACC | + |
| 8 | AGCCGGTGTTAGTAAGAAAT | + |
| ... | ... | ... |
For non-directional sequencing library, we need to know the sequence vector without sgRNA to determine the strand of extracted sgRNA. The sequence should be split into two end at the insertion site of sgRNA, one end should be named as 5 and another is 3.
>5
ccagaGagggcctatttcccatgattccttcatatttgcatatacgatacaaggctgttagagagataattagaattaatttgactgtaaacacaaagatattagtacaaaatacgtgacgtagaaagtaataatttcttgggtagtttgcagttttaaaattatgttttaaaatggactatcatatgcttaccgtaacttgaaagtatttcgatttcttggctttatatatcttgtggaaaggacgaaacaccg
>3
gttttagagctagaaatagcaagttaaaataaggctagtccgttatcaacttgaaaaagtggcaccgagtcg
Extracted sgRNA for each sample and an HTML report for quality control.
The HTML report was powered by Multiqc, see the example in output directory.
- This script only suits pair-end non-directional sequencing data and the insertion site of sgRNA is near 3' end.
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