Perturb-seq Guide Barcodes (GBC) Library
(Pooled Library #85968)
Barcode library for cloning gRNA sublibraries for single cell CRISPR analysis.
- Vector Backbone
- Depositing Labs
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This library is delivered as suspended DNA in a microcentrifuge tube on blue ice. The tube's contents will not necessarily be frozen. For best results, minimize freeze-thaw cycles.
Terms and Licenses
The Perturb-seq GBC library (also referred to as pBA571) is intended for cloning of individual protospacer sequences into the sgRNA expression cassette followed by isolation and verification of single barcoded expression vectors, as in the paper. It has not been optimized for pooled cloning. Of note, restriction digests indicate the presence of spurious DNA introduced during library construction from pBA439 that has not been observed to carry over into isolated sgRNA expression derivatives.
For amplification of the library, the use of high efficiency, electrocompetent cells is strongly advised. To ensure the maintenance of library diversity, calculate your transformation efficiency. 30x coverage of the library is recommended.
This library has been verified using individually cloned guide RNAs. Guide RNA protospacer sequences were individually cloned into the BstXI and BlpI sites by direct ligation. Protospacers and corresponding GBCs were then verified by sequencing. Final Perturb-seq vectors containing barcodes that introduce the conserved polyadenylation signal AATAAA should be discarded.
This pooled library was created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the pooled library was described, and include Addgene in the Materials and Methods of your future publications.
Example for your Materials & Methods section:Perturb-seq GBC library was a gift from Jonathan Weissman (Addgene ID #85968)
For your References section:A Multiplexed Single-Cell CRISPR Screening Platform Enables Systematic Dissection of the Unfolded Protein Response.. Adamson B, Norman TM, Jost M, Cho MY, Nunez JK, Chen Y, Villalta JE, Gilbert LA, Horlbeck MA, Hein MY, Pak RA, Gray AN, Gross CA, Dixit A, Parnas O, Regev A, Weissman JS. Cell. 2016 Dec 15;167(7):1867-1882.e21. doi: 10.1016/j.cell.2016.11.048 PubMed 27984733