CHyMErA Knockout Libraries
(Pooled Library #155199, #155200)

• Purpose:

  The Cas Hybrid for Multiplexed Editing and screening Applications (CHyMErA) lentiviral libraries express hybrid Cas9-Cas12a guide RNAs under a single U6 promoter.
  
  The Paralog & Dual-targeting hybrid guide RNA Library targets human paralog pairs with single- and combinatorial-targeting hgRNAs and can be used to identify genetic interactions between paralogs. The library also contains single- and dual-targeting hybrid guides that target ~5,000 highly expressed genes permitting capture of gene ablation phenotypes with increased sensitivity.
  
  The Exon-deletion hgRNA Library deletes human alternative cassette exons and disrupts genes containing alternative exons, allowing investigation into alternative exon function.

• Vector Backbone:

  pLCHKO - does not express Cas9 or Cas12a

• Depositing Labs: Jason Moffat
  Benjamin Blencowe
• Publication: Gonatopoulos-Pournatzis et al Nat Biotechnol. 2020 May;38(5):638-648. doi: 10.1038/s41587-020-0437-z. Epub 2020 Mar 16.

Library Details

• Species: Human
• Genes targeted:

  
  Paralog & Dual-targeting library: 6,333 genes (1,344 paralogs & 4,993 genes)
  Exon-deletion library: 2,157 alternative cassette exons in 1,745 genes

• hgRNAs:

  
  Paralog & Dual-targeting library: 92,746
  Exon-deletion library: 91,948

• Controls:

  
  Paralog & Dual-targeting library: 4,993 intergenic and non-targeting hgRNAs
  Exon-deletion library: 1,647 intergenic control hgRNAs

• Lentiviral Generation: 3rd

Library Shipping

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-thaws.

• Volume: ∼20 µL
• Concentration: 50 ng/µL

Resource Information

• Protocols:
  • Library Transformation and NGS Protocols (DOCX, 22 KB)
• Depositor Data:
  • Paralog & Dual-targeting hgRNA sequences (7.0 MB)
  • Exon-deletion hgRNA sequences (6.1 MB)
  • Distribution of NGS Reads- Paralog & Dual-targeting Library (135 KB)
  • Distribution of NGS Reads- Exon-deletion Library (134 KB)
• Terms and Licenses:
  • UBMTA

Depositor Comments

Screening analysis is described in:
Pooled Lentiviral CRISPR-Cas9 Screens for Functional Genomics in Mammalian Cells. Aregger M, Chandrashekhar M, Tong AHY, Chan K, Moffat J Methods Mol Biol. 2019;1869:169-188. doi: 10.1007/978-1-4939-8805-1_15. PMID 30324523

The depositing lab has provided analysis algorithms and R code in their Github repositories:

CHyMERa Dual-targeting Scoring
CHyMERa Exon-deletion Scoring

Read depositor Jason Moffat's description of CHyMErA in this Nature Bioengineering Behind The Paper article.