MLP Retroviral shRNA-mir focused setsPLATINUM Select MLP Retroviral shRNA-mir sets targeting popular gene families and pathways are available and custom arrays can be created. These focused sets are available in a 96 well arrayed (bacterial glycerol stock) and pooled formats making them ideal for RNAi screening for a variety of biological assays. The flexibility of formats and the renewable nature of vector-based RNAi provide economical library options for screening or as a long-term resource suiting the needs of most RNAi applications.
Focused sets were created using resources from the National Institute of Health’s Cancer Genome Anatomy Project (CGAP) and Gene Ontology (GO). Annotation is based on electronic and manual literature-based curation as well as sequence analysis. (See Annotation Standard Operating Procedures from the Gene Ontology Consortium for more information.)
Benefits of the arrayed libraries include:
Don’t see your gene family or pathway of interest? Contact us at firstname.lastname@example.org to ask about custom arrayed or pooled shRNA-mir sets.
pMLP Vector DesignThe shRNA-mir sequences have been cloned in to the pMLP vector which has the following characteristics:
Arrayed and pooled screening with shRNA-mirVector-based RNAi screens can be implemented using two different strategies. The classic arrayed strategy, adapted from the small molecule screening field, is a well-by-well screen testing one gene per well using plasmid DNA or viral particle formats. A number of possible readouts have proven successful with this technique including cell viability, reporter assays, and morphological changes. Additionally, high-content screening with RNAi is a popular way to perform phenotypic screens on a large number of genes in parallel (Hu, G and Luo Ji 2012).
The multiplexed (pooled) strategy enables hundreds to thousands of genes to be simultaneously targeted. This is only possible using viral vector-based RNAi. Multiple constructs are pooled together, packaged into virus and cells are transduced at a multiplicity of infection (MOI) ensuring single integrations per cell. From this, each transduced cell becomes its own experiment and positive or negative selection can be used to determine hits (see figure 1). shRNA-mir that induce or diminish a phenotype can be detected using next generation sequence analysis of the shRNA integrated into the target cell’s genome.
Hu, G and Luo, Ji (2012) A primer on using pooled shRNA libraries for functional genomic screens. Acta Biochim Biophys Sin 2012, 44: 103–112.
The schematic to the right depicts this process.