Mike,

Finally I figured out  the problem. It is not a bug. It is the searching criteria  max.mismatch =5 and filtering criteria min.score = 0.5. If you set max.mismatch = 5 and min.score = 0.1. The summary file will be there. What happened is that with your  initial parameter setting, there is no off-target found. Maybe I should have the function to print out “No off target found with your searching and filtering criteria”.

Best regards,

Julie



On 8/4/14 11:03 AM, "Brodsky, Michael" <Michael.Brodsky@umassmed.edu> wrote:

Hi. I tried two more tests.

First, I ran the same script on Rstudio on the cluster. The cluster packages are all the standard (not development) version of R and do not give the Warning message for R version 3.1.1. So, it is NOT due to using a different version of R for the packages.

Second, there is only one target site in the fasta file, so I also ran it with find gRNA = TRUE. This gave the same problem where the summary file does not list any gRNAs.

Mike




On Aug 4, 2014, at 10:50 AM, Michael Brodsky wrote:

Hi Julie,

Here is the first problem. I ran an off target analysis with existing target site (find gRNA = FALSE). I got a good off target file, but the summary file is blank. I have attached the input file and the command set. The session info is below. I got a warning that I used a more recent (developer?) version of R to construct the packages. Do I need to uninstall and reinstall these?

thanks,

mike



Warning messages:
1: package ‘CRISPRseek’ was built under R version 3.1.1
2: package ‘Biostrings’ was built under R version 3.1.1
3: package ‘IRanges’ was built under R version 3.1.1
4: package ‘GenomeInfoDb’ was built under R version 3.1.1
5: package ‘GenomicRanges’ was built under R version 3.1.1

Warning messages:
1: package ‘GenomicFeatures’ was built under R version 3.1.1
2: package ‘AnnotationDbi’ was built under R version 3.1.1



R version 3.1.0 (2014-04-10) -- "Spring Dance"
Copyright (C) 2014 The R Foundation for Statistical Computing
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Type 'license()' or 'licence()' for distribution details.

  Natural language support but running in an English locale

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Type 'contributors()' for more information and
'citation()' on how to cite R or R packages in publications.

Type 'demo()' for some demos, 'help()' for on-line help, or
'help.start()' for an HTML browser interface to help.
Type 'q()' to quit R.

[R.app GUI 1.63 (6734) x86_64-apple-darwin10.8.0]

[Workspace restored from /Users/brodsky/.RData]
[History restored from /Users/brodsky/.Rapp.history]

> getwd()
[1] "/Users/brodsky/Desktop/current/Nuclease_projects/inProgress/Freeman_tobi1/tobi2_test"
> library(CRISPRseek)
Loading required package: BiocGenerics
Loading required package: parallel

Attaching package: ‘BiocGenerics’

The following objects are masked from ‘package:parallel’:

    clusterApply, clusterApplyLB, clusterCall, clusterEvalQ, clusterExport, clusterMap, parApply, parCapply, parLapply,
    parLapplyLB, parRapply, parSapply, parSapplyLB

The following object is masked from ‘package:stats’:

    xtabs

The following objects are masked from ‘package:base’:

    anyDuplicated, append, as.data.frame, as.vector, cbind, colnames, do.call, duplicated, eval, evalq, Filter, Find, get,
    intersect, is.unsorted, lapply, Map, mapply, match, mget, order, paste, pmax, pmax.int, pmin, pmin.int, Position,
    rank, rbind, Reduce, rep.int, rownames, sapply, setdiff, sort, table, tapply, union, unique, unlist

Loading required package: Biostrings
Loading required package: S4Vectors
Loading required package: IRanges
Loading required package: XVector
Loading required package: BSgenome
Loading required package: GenomeInfoDb
Loading required package: GenomicRanges
Loading required package: rtracklayer
Warning messages:
1: package ‘CRISPRseek’ was built under R version 3.1.1
2: package ‘Biostrings’ was built under R version 3.1.1
3: package ‘IRanges’ was built under R version 3.1.1
4: package ‘GenomeInfoDb’ was built under R version 3.1.1
5: package ‘GenomicRanges’ was built under R version 3.1.1
> library("BSgenome.Dmelanogaster.UCSC.dm3")
> library("TxDb.Dmelanogaster.UCSC.dm3.ensGene")
Loading required package: GenomicFeatures
Loading required package: AnnotationDbi
Loading required package: Biobase
Welcome to Bioconductor

    Vignettes contain introductory material; view with 'browseVignettes()'. To cite Bioconductor, see
    'citation("Biobase")', and for packages 'citation("pkgname")'.


Attaching package: ‘AnnotationDbi’

The following object is masked from ‘package:BSgenome’:

    species

The following object is masked from ‘package:GenomeInfoDb’:

    species

Warning messages:
1: package ‘GenomicFeatures’ was built under R version 3.1.1
2: package ‘AnnotationDbi’ was built under R version 3.1.1
> outputDir <- getwd()
> inputFilePath <- "tobi2.fa"
> REpatternFile <- system.file("extdata", "NEBenzymes.fa", package = "CRISPRseek")
>
> offTargetAnalysis(inputFilePath, findgRNAs = FALSE, findgRNAsWithREcutOnly = FALSE, REpatternFile = REpatternFile, findPairedgRNAOnly = FALSE, BSgenomeName = Dmelanogaster, txdb = TxDb.Dmelanogaster.UCSC.dm3.ensGene, max.mismatch = 3, chromToSearch = "all", outputDir = outputDir, overwrite = TRUE)
Validating input ...
>>> Finding all hits in sequences chr2L ...
>>> DONE searching
>>> Finding all hits in sequences chr2R ...
>>> DONE searching
>>> Finding all hits in sequences chr3L ...
>>> DONE searching
>>> Finding all hits in sequences chr3R ...
>>> DONE searching
>>> Finding all hits in sequences chr4 ...
>>> DONE searching
>>> Finding all hits in sequences chrX ...
>>> DONE searching
>>> Finding all hits in sequences chrU ...
>>> DONE searching
>>> Finding all hits in sequences chrM ...
>>> DONE searching
>>> Finding all hits in sequences chr2LHet ...
>>> DONE searching
>>> Finding all hits in sequences chr2RHet ...
>>> DONE searching
>>> Finding all hits in sequences chr3LHet ...
>>> DONE searching
>>> Finding all hits in sequences chr3RHet ...
>>> DONE searching
>>> Finding all hits in sequences chrXHet ...
>>> DONE searching
>>> Finding all hits in sequences chrYHet ...
>>> DONE searching
>>> Finding all hits in sequences chrUextra ...
>>> DONE searching
Building feature vectors for scoring ...
Calculating scores ...
Annotating, filtering and generating reports ...
Done. Please check output files in directory  /Users/brodsky/Desktop/current/Nuclease_projects/inProgress/Freeman_tobi1/tobi2_test/
>
<tobi2_rscript.txt><tobi2.fa>



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