Human 87K Knockout Library (UbiC-Puro) (packaged, large scale) – academia

Description

Pooled Lentiviral CRISPR KO sgRNA Library for Human Genome Wide CRISPR KO Screenings

• Genome-wide coverage with a single pooled library featuring ~87,000 constructs – download gene list here
• 19,382 human protein-coding genes covered by 4 CRISPR Knockout sgRNAs each, targeting functional domains and conserved transcript region
• Gene annotations based on the latest reference genome GRCh38
• 1,859 human microRNAs covered by up to 4 CRISPR Knockout sgRNAs
• Controls included: 1,000 non-targeting control sgRNAs; 400 intron-targeting sgRNAs against the introns of 100 genes; 400 intergenic sgRNAs, and 400 sgRNAs targeting 2 Safe Harbor sites (AAVS1 and CLYBL) are additionally included in the library
• sgRNA design based on Broad Institute´s latest design rules to ensure highest knockout and lowest off-target efficiencies
• HECG-modified tracrRNA in the sgRNA scaffold enables enhanced Cas9 binding and sgRNA efficacy
• NGS-verified uniquely tight sgRNA distribution in the library pool which applies to all Cellecta sgRNA libraries

Side-by-side comparisons of Cellecta genome-wide sgRNA libraries with two other suppliers of genome wide sgRNA libraries

Human Genome Wide CRISPR KO Library – pooled lentiviral sgRNA library with Puro marker

Cellecta’s single-module Human Genome-Wide CRISPR KO Library cloned in the pRSGUP-U6-sg-UbiC-Puro vector was designed to be small enough (~87,000 constructs covering 19,382 protein coding genes and 1,859 miRNAs) to enable convenient genome-wide dropout viability or gain of function screens. With up to 4 sgRNAs targeting each miRNA and 4 sgRNAs targeting each human protein coding gene gene – designed to target functional domains and regions conserved between alternatively spliced transcripts – the library has deep gene coverage generating efficient results.

All guide designs are using Broad Institute´s latest design rules – CRISPick, Drepanos et al., 2025 – for optimal on target efficiency and lowest off target efficiency, as well as Cellecta´s in house algorithms for knock out efficiency ranking, to achieve most robust knockout screening results. Furthermore the tracrRNA portion of the sgRNA scaffold features the HECG design for further sgRNA efficacy improvement upon the previous HEAT design (DeWeirdt et al.:  Accounting for small variations in the tracrRNA sequence improves sgRNA activity predictions for CRISPR screening)

Deliverables:

• 1 x 10e9 TU ready-to-transduce VSV-G typed pseudolentiviral particles, functionally titered in HEK293 cells. Particles have been purified and concentrated by LentiFuge Concentration Reagent and are delivered dissolved in PBS.
• Sequence File featuring all sgRNA sequences and their distribution in the pool