CRISPRAnalyzeR run stalling at 91%

[Kartik Chandran]

Dear Jan,

I am a new user, so my apologies if I'm doing anything obviously silly. I am trying to analyze a MiSeq run of the Gecko V2 library we have created. I uploaded two fastq.gz files for analysis, along with the <FASTA_GeckoV2_all.fasta> library file you provided. I used the standard LentiCrispr regular expression to extract the gRNA sequences, which seems to check out fine. The run starts OK but repeatedly stalls at 91%. I have provided below some lines from my fastq file. 

My setup:

Mac running OS X Yosemite (10.10.5) 

Docker 17.03.1-ce-mac12 (17661)

Kitematic 0.13.1

Thanks for your help!

Kartik

+
CCCCCGGGGGEFC@C@FGGGGE@GG78FG<FCGGGGF:;FGGGCFCFGE8FGGGEFECF,<,CDC<C,CFGGGD@6@B@8FEACEG<EG=F?FFC,6C<<
@M01243:170:000000000-B5VFP:1:1101:15920:1112 1:N:0:NTCACG
GATGCACATCTGCTTTATATATCTTGTGGAAAGGACGAAACACCGATTTCTTGCAGGTCCAACATGTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCT
+
CCCCCGGGGGGCGGGGGGGGGAFGGGGGGF,C,CFGGGGGGGFG@:CFFGGGFGCFFGCECFDDFFFG9FFFCEGAE<FGGGGGGGFGFGGGFGGGGFF?F
@M01243:170:000000000-B5VFP:1:1101:19683:1112 1:N:0:NTCACG
CGATTGCTCGACGCTTTATATATCTTGTGGAAAGGACGAAACACCGGCTCTTCTGAGATCTAAGTTGTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC
+
CCCCCGGGGDGC@CFGFGGC<E<EFFEFGGGGGGCGGCEEE7<FGGGGGGGGGGGAA,CFGGAFFFGGFGGGGFF9CCFFFEFDDCFFFFFC6,6C<CEFE
@M01243:170:000000000-B5VFP:1:1101:20341:1112 1:N:0:NTCACG
CGATCGATTTGAGCCTGCTTTATATATCTTGTGGAAAGGACGAAACACCGCGCCAAGTCATCCGACCGTGTTTTAGAGCTAGAAATAGCAAGTTAAAATA
+
-ABBC:FEFGGGEGGFGGGFGGGGGGGGGGGGGGGGGGCDFGGGGGGGGDFF+:FF@FGG9CECGGGEGGGGGGGGEFCCEGFGFFGFDGFGGFCFFEFG
@M01243:170:000000000-B5VFP:1:1101:14359:1112 1:N:0:NTCACG
TAAGTAGAGGCTTTATATATCTTGTGGAAAGGACGAAACACCGGCAGGAACTGACCGCAAACGGTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCT

[Jan Winter]

Hi Kartik, sorry for my late response.

We had some issues with the Gecko library before that should have been fixed... at least I thought so (I don't have Gecko data here for testing).

I guess it stalls during the FASTQ extraction? Or is it the analysis?

If it is the analysis, it could be ScreenBEAM analysis which takes some time to calculate.

Could you please do me favor so that I can nail the issue down:

- get the latest BETA using docker pull boutroslab/crispranalyzer:1.16BETA

- do the FASTQ extraction again

- wait until it stalls (91-95 % can take some time since it generates a FASTQ quality report)

- once it does not go on for 30 minutes or so, please proceed to the About CRISPRAnalyzeR page and hit the "Download Log Files" button.

Please send me the log file archive to

jan.w...@dkfz-heidelberg.de

Then I can have a closer look what might have been the issue and I am sure we will figure it out :)

Best and thanks for your help

Jan