shERWOOD-UltramiR Human Genome Wide Arrayed shRNA Library

Description

Human Genome Wide Arrayed shRNA Library – Premade shRNA Library for Arrayed RNAi Screens

Your shRNA Resource for Individual, Combinatorial, or Multiplexed Gene Knockdown at genome-wide scale

Targets over 18,000 human genes with an average of 6 shRNA constructs per gene, resulting in approximately 108000 shRNA constructs, arrayed as glycerol stock clones in 1125  96-well plates

shERWOOD shRNA design + Optimized shRNA processing = Superior Knockdown

• Every shRNA construct has been 100% sequence-verified
• The proprietary shERWOOD algorithm was used to design all shRNA
• The UltramiR microRNA scaffold increases shRNA processing and potency
• High efficiency retroviral vector backbone for high-titer viral particle production

This human genome shRNA construct collection was created in collaboration with Cold Spring Harbor Laboratory (Knott et al., Molecular Cell) using the shERWOOD algorithm for shRNA design and embedding the shRNA sequences in the UltramiR scaffold to enable high efficiency gene-by-gene/well-by-well, & combinatorial, or multiplexed gene knockdown screens by looking at molecular phenotypes.

The library uses an alternate microRNA scaffold called “UltramiR“. The UltramiR scaffold has been optimized for increased shRNA processing and potency based on the key determinants for primary microRNA processing (Auyeung et al 2013)

The shERWOOD algorithm is based on the functional testing of over 250,000 shRNA sequences using a high-throughput sensor assay and uses key sequence characteristics for predicting shRNA potency to select the rare shRNA designs that are potent at single copy representation in the genome:

Construction and Validation of an shRNA-Specific Predictive Algorithm (hhERWOOD) (A) Consolidated cross-validation of predictions versus sensor scores for all shRNAs in the Fellmann et al. (2011) data set (shRNAs are separated by the guide 5 0 nucleotide). (B) GO term instances associated with the targeted gene set selected for shRNA validation screens. (C) GO term instances associated with genes for which at least two hairpins were significantly depleted in each of the TRC, Hannon-Elledge (HE), and shERWOOD (SW) validation screens. (D) The percentage of shRNAs targeting consensus-essential genes that were depleted in each of the TRC, HE, and shERWOOD shRNA screens. The plot was made with the Matlab Boxplot function using default parameters. The edges of the box are the 25 th and 75 th percentiles. The error bars extend to the values q3 + w(q3 ? q1) and q1 ? w(q3 ? q1), where w is 1.5 and q1 and q3 are the 25 th and 75 th percentiles. (E) Average log-fold change for shRNAs targeting consensus-essential genes (per gene) for each of the TRC, EH, and shERWOOD validation screens. The plot was made with the Matlab Boxplot function using default parameters. The edges of the box are the 25 th and 75 th percentiles. The error bars extend to the values q3 + w(q3 ? q1) and q1 ? w(q3 ? q1), where w is 1.5 and q1 and q3 are the 25 th and 75 th percentiles. (F) The percentage of shRNAs corresponding to consensus-essential genes that, for any given shERWOOD score, were depleted in the shERWOOD validation screen.

The shRNAs are cloned into a retroviral vector that contains a Neomycin selection marker and a ZsGreen fluorescent reporter:

Retroviral shRNA vector pLMN-ZsGreen-Neomycin

CONTENTS:

The  shERWOOD-UltramiR Human Genome Wide Arrayed shRNA Library is delivered as barcoded 96-well glycerol stock plates – 1125 plates total. Plates are sealed with aluminum sealing tape and are shipped on dry ice.

The plates should be stored at -80C upon arrival.