Bikard E. coli Genome-wide Inhibition Library
(Pooled Library #115927)

• Purpose:

  This library contains ~93,000 unique guide RNAs targeting random positions along the genome of E. coli MG1655 with a NGG PAM motif. The library contains an average of 19 targets per gene and was made by cloning a pool of guide RNAs oligos into psgRNA (#114005) (ori: ColE1) by Gibson assembly.

  This library can be used with SpdCas9 to perform pooled CRISPRi screens as shown ( Cui et al., 2018 ; Rousset et al. 2018). Screens can be performed using strain LC-E75 (#115925), E. coli strain MG1655 expressing dCas9 under the control of a Tet promoter integrated at the 186 attB site, or strain FR-E01 (#118727), E. coli strain MG1655 expressing dCas9 under the control of a Tet promoter integrated at the HK022 attB site.

• Vector Backbone:

  psgRNA - does not express Cas9

• Depositing Labs: David Bikard
• Publication: Cui et al. Nat Commun. 2018 May 15;9(1):1912. doi: 10.1038/s41467-018-04209-5.

Library Details

• Species: Escherichia coli MG1655
• Genes targeted: ~4,500
• gRNAs: 92,919
• Controls: 1 - backbone is used as a control

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.

NOTE: 100 ng of the library can be electroporated into cells for an experiment. Sufficient sample is provided for several experiments and amplification of the library.

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

Resource Information

• Protocols:
  • Library Transformation Protocol (17 KB)
  • Illumina Screening Protocol (15 KB)
  • Illumina Sequencing Protocol (20 KB)
• Depositor Data:
  • List of gRNAs (7.7 MB)
  • NGS Read Counts (3.4 MB)
• Terms and Licenses:
  • UBMTA

Depositor Comments

The depositing lab recommends using their code for analyzing Illumina sequencing results.

For additional information on use of this library, please see Rousset et al., 2018.