Extensive human RNAi library covers over 80% of the genes within key categories such as cell cycle, GPCRs, proteolysis and transcription
MRC Geneservice has added to its highly successful RNA interference libraries with a new extensive human RNAi library. Over 9000 human genes are targeted with 18,000 shRNA (short-hairpin RNA) plasmid constructs.
This covers over 80% of the genes within key categories such as cell cycle, GPCRs, proteolysis and transcription.
Using the short-hairpin RNAi concept, this library provides researchers with an advanced large-scale tool for biological discovery.
The hairpin sequences, constructs and protocols have all been carefully developed and tested by Greg Hannons' laboratory at the Cold Spring Harbor Laboratories (CSHL), New York.
Many essential genes are targeted with two or more shRNA constructs.
Moreover, shRNA fate can be tracked using DNA 'bar codes' unique to each construct. The constructs have been designed and cloned into a specialised retroviral vector, which allows the human library to utilise RNA polymerase III promoters to generate more stable expression of shRNA.
This also avoids the expense of chemically synthesised short interfering RNAs (siRNAs).
MRC Geneservice's human RNAi system utilises many recent advances in RNAi, such as shRNA cassette transfer via homologous recombination, which allows greater control over positive selection.
The hairpins themselves are relatively long at 29 nucleotides (nt) of dsRNA and have a simple four nucleotide loop.
The cassette also contains the U6 small nuclear RNA promoter with a 27nt leader sequence and the human miR30 microRNA loop and flanking region.
These additions give greater expression and significantly increase Drosha and Dicer processing.
Moreover, only coding sequences are targeted by the shRNA constructs and there are at least three mismatches to any other gene.
As well as this, selected targets, where possible, have sequence identity with the mouse homologue, allowing cross species use.
This powerful research tool complements and expands MRC Geneservice's RNAi Library resources, which also includes extensive libraries for Drosophila melanogaster and Caenorhabditis elegans gene knockdowns.
MRC Geneservice can now provide researchers with the ability to rapidly screen for the function of specific genes and search for novel therapeutic targets across a range of genomes.
In addition to RNAi libraries, MRC geneservice offers an array of other tools for researchers including genomic, cDNA, RNA and proteomic tools, as well as contract services for DNA and RNA extraction and processing.
