FYI, to search for gRNAs without performing off target analysis, please set chromToSearch = "" and there is no need to set the BSgenomeName https://www.bioconductor.org/packages/devel/bioc/vignettes/CRISPRseek/inst/doc/CRISPRseek.pdf
(2.6 Scenario 6. Quick gRNA finding without target or off-target analysis).
To perform off target analysis, you do need to set BSgenomeName. Here is the instruction on how to forge a BSgenome
If it is too much work to forge a genome, you could try the function offTargetAnalysisOfPeakRegions (http://www.bioconductor.org/packages/release/bioc/manuals/GUIDEseq/man/GUIDEseq.pdf
) in the GUIDEseq package http://www.bioconductor.org/packages/release/bioc/html/GUIDEseq.html
Here is an example assuming gRNAFile contains the gRNA(s) in fasta format and fastaSequenceFile contains the genome sequence of your interest in fasta format.
offTargets <- offTargetAnalysisOfPeakRegions(gRNA = gRNAFile, peaks = fastaSequenceFile, format="fasta", peaks.withHeader = FALSE, upstream = 0, downstream =0, PAM.size = 3, gRNA.size = 20, PAM = "NGG", PAM.pattern = "N[A|G]G$", max.mismatch = 3, outputDir = outputDir, orderOfftargetsBy = "predicted_cleavage_score", allowed.mismatch.PAM = 1, overwrite = TRUE )
From: "millepattes.intersites [bioc]" <firstname.lastname@example.org<mailto:email@example.com>>
Reply-To: "firstname.lastname@example.org<mailto:email@example.com>" <firstname.lastname@example.org<mailto:email@example.com>>
Date: Tuesday, March 14, 2017 3:27 PM
To: Lihua Julie Zhu <firstname.lastname@example.org<mailto:email@example.com>>
Subject: [bioc] CRISPRseek with genome not available in BSgenome
Activity on a post you are following on support.bioconductor.org<https: support.bioconductor.org="">
User millepattes.intersites<https: support.bioconductor.org="" u="" 12592=""/> wrote Question: CRISPRseek with genome not available in BSgenome<https: support.bioconductor.org="" p="" 93835=""/>:
I would like to use CRISPRSeek to design gRNA targeting a genome which is not available in BSgenome database. This concerns the bacterial genome of Streptomyces ambofaciens ATCC 23877 (NCBI Reference Sequence: NZ_CP012382.1).
Is there any tutorial explaining how to use CRISPRSeek in this case. I found some indications in a supplementary information (article published in PloSOne, 2014), but I did not clearly understand how to create the file (with the genome, and with the target sequence) as well as how I should modify the instructions.
I thank you very much for your help.
Post tags: bsgenome, crisprseek
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