I have Human CRISPR Knockout Pooled Library (GeCKO v2) from Addgene ([https://www.addgene.org/pooled-library/zhang-human-gecko-v2/])[2] which I have amplified according to the reference paper -
- Joung, J., Konermann, S., Gootenberg, J. et al. Genome-scale CRISPR-Cas9 knockout and transcriptional activation screening. Nat Protoc 12, 828–863 (2017). https://doi.org/10.1038/nprot.2017.016
- Improved vectors and genome-wide libraries for CRISPR screening . Sanjana NE, Shalem O, Zhang F. Nat Methods. 2014 Aug;11(8):783-4. doi: 10.1038/nmeth.3047. 10.1038/nmeth.3047PubMed 25075903
I had sent it for NGS so that the distribution of the sgRNAs could be validated. only one half of the library i.e. Va could be demultiplexed while the other half of the library could not be demultiplexed even though the index sequences that I used were used were correct and matched with the sample sheet given in Illumina Adapter Sequences. yet I was not able to demultiplex the data. moreover the first half of the library that I was able to demultiplex had a very poor read2 quality. I would really appreciate if anyone who has validated the GeCKO library, would be able to help me.
