BCH709 Introduction to Bioinformatics: CRISPR/Cas9 database build

I. scapularis genome for CRISPR/Cas9

Convert your genome file to ChopChop format Requested by Dr. Monika Gulia-Nuss and Dr. Arvind Sharma

Gulia-Nuss Lab.

Dr. Monika Gulia-Nuss Figure 1. Jeremiah Reyes (Grad special), Preston, Dr. Arvind Sharma, Dr. Monika Gulia-Nuss and Dr. Andrew Nuss.

Ticks Figure 2. Genes associated with the unique parasitic lifestyle of Ixodes scapularis. Genomic insights into the Ixodes scapularis tick vector of Lyme disease

ChopChop

http://chopchop.cbu.uib.no/

Genome submission instructions

https://chopchop.cbu.uib.no/submissions

Citations for ChopChop

Kornel Labun; Tessa G. Montague; James A. Gagnon; Summer B. Thyme; Eivind Valen. (2016). CHOPCHOP v2: a web tool for the next generation of CRISPR genome engineering. Nucleic Acids Research; doi:10.1093/nar/gkw398

Tessa G. Montague; Jose M. Cruz; James A. Gagnon; George M. Church; Eivind Valen. (2014). CHOPCHOP: a CRISPR/Cas9 and TALEN web tool for genome editing. Nucleic Acids Res. 42. W401-W407

Requirement

Any linux system or Any terminal Web browser with internet Your passion

Processing

1. Download Tick genome from Vectorbase

Download links

Prepare following files.

Gene feature file (GFF)

Ixodes-scapularis-Wikel_BASEFEATURES_IscaW1.6.gff3.gz

Genome sequence (scaffold)

Ixodes-scapularis-Wikel_SCAFFOLDS_IscaW1.fa.gz

Download

$ wget https://www.vectorbase.org/sites/default/files/ftp/vbo_archive_20180731_0.zip

Unzip data

$ unzip vbo_archive_20180731_0.zip

Check the files use “space bar” and quit with “q” for “less”

$ zcat  Ixodes-scapularis-Wikel_SCAFFOLDS_IscaW1.fa.gz | less 
$ zcat Ixodes-scapularis-Wikel_BASEFEATURES_IscaW1.6.gff3.gz | less

*Check the number of scaffold in GFF and scaffold(fa)

$ zcat Ixodes-scapularis-Wikel_BASEFEATURES_IscaW1.6.gff3.gz | egrep -c  "sequence-region"
$ zcat  Ixodes-scapularis-Wikel_SCAFFOLDS_IscaW1.fa.gz | egrep -c ">"

*Result (number of scaffolds in GFF and fasta files)

$ 369492

2. Prepare gff3ToGenePred.

gff3ToGenePred is validating and GFF file format to Gp (GenePred) format

Download gff3ToGenePred

$ wget http://hgdownload.cse.ucsc.edu/admin/exe/linux.x86_64/gff3ToGenePred

Check the excute

$ chmod 775 ./gff3ToGenePred      
$ ./gff3ToGenePred

3. Convert gff3 to Gp

Convert with gff3ToGenePred

$ ./gff3ToGenePred Ixodes-scapularis-Wikel_BASEFEATURES_IscaW1.6.gff3.gz Ixodes-scapularis-Wikel_BASEFEATURES_IscaW1.pred -bad=Ixodes-scapularis.bad

Check the BAD file (Should be nothing)

$ less Ixodes-scapularis.bad

Check the Gp file (Should include exon locations, gene name and scaffold ID)

$ less Ixodes-scapularis-Wikel_BASEFEATURES_IscaW1.pred

4. Prepare the file for ChopChop

$ mkdir Ixodes-scapularis_ChopChop ## make directory
$ mv Ixodes-scapularis-Wikel_BASEFEATURES_IscaW1.pred Ixodes-scapularis_ChopChop ## move files
$ mv Ixodes-scapularis-Wikel_SCAFFOLDS_IscaW1.fa.gz Ixodes-scapularis_ChopChop ## move files
$ tar -cvzf Ixodes-scapularis_ChopChop.tar.gz Ixodes-scapularis_ChopChop ## compress
$ ls -lh Ixodes-scapularis_ChopChop.tar.gz

5. Contact to ChopChop

The file size is around 453Mb after compress. We need to use BOX to share this files. I prefer to share it by link. How to share it by link

Send email to Tessa G. Montague

tessa.chopchop@gmail.com

Take home message

  1. what is “’ ’” ?
  2. What is “zcat” ?
  3. How to compress and decompress files? explain option (ex cvf, xzvf)
  4. Why do we need to “Check the number of scaffold in GFF and scaffold” ?
  5. Explain GFF
  6. Can you do same thing in Anopheles stephensi?
  7. Explain about CHOPCHOP statistical approach