GeneNet™ Human Druggable gene siRNA Library

  • 43,800 siRNA sequences targeting 8,500 named human genes

  • Selectable puromycin resistance allows positive identification and selection of transduced cells

  • Pre-packaged, ready-to-use format—transduce your cells the day you receive the library

  • High-throughput identification of functional siRNAs by Affymetrix GeneChip® hybridization

  • Also available in plasmid form, you can now produce large quantities of packaged siRNA library in your own lab

Data comparing hybridization of amplified and labeled siRNA templates prepared from packaged viral particles and from genomic DNA extracted from target cells transduced with the library.  The data demonstrates that the transduction of the library is highly efficient and does not bias the library with respect to representation.

The GeneNet™ Human Druggable Genome siRNA Library contains 43,800 siRNA templates targeted to 8,500 well-characterized human genes listed in the NCBI RefSeq database. For most of the target genes, five different siRNA sequences were designed using our proprietary algorithm.  The library is cloned into the FIV-based pSIF1-H1-Puro shRNA Expression Vector, which confers resistance to the antibiotic puromycin.  This single-promoter siRNA vector expresses cloned short-hairpin RNA (shRNA) from a single H1 RNA polymerase III promoter.

The library is available in both pre-packaged and plasmid form. The pre-packaged VSV-G pseudotyped viral stock can be used to transduce your cells of choice immediately upon receipt, or be stored at -70oC for several months. With the library in plasmid form, you can perform the pseudoviral packaging step in your own cell culture facility. Enough plasmid is provided to yield about 109 ifu of packaged siRNA library.

As described in the GeneNet Libraries “How it Works” section, after transducing the library into your cells of interest, you select those cells that display the desired phenotype or response using whatever technique is appropriate for your system. Following selection, the lentiviral inserts—containing the siRNA templates—are easily recovered by PCR and identified using the Human Genome Focus GeneChip® Array manufactured by Affymetrix (Santa Clara, CA). The siRNA template sequences were designed to hybridize to oligonucleotide probes on this array to facilitate identification. Although more tedious, you can also use sequencing to identify the particular siRNA sequences in selected cells.

To confirm representation, siRNA templates for the library were packaged in viral particles, amplified, labeled, and hybridized to the Affymetrix GeneChip®. Similarly, siRNA templates were amplified from the genomic DNA extracted from target cells transduced with the library. These templates were also labeled and hybridized to the GeneChip®. The hybridization results for the Human Druggable Genome from both arrays are compared in the graph shown above. Overall, in both the viral- and cell-derived samples, more than 55% of the gene-specific siRNA template species used in construction of the library were easily detectable above background. In addition, the correlation of the relative amounts of different siRNA template species between both samples was very high, indicating that there is not appreciable loss or change in representation with transduction. These results indicate that the library delivers a representative number of siRNA sequences that target the majority of genes for which the library was originally designed. Also, the high correlation between the pre- and post-transduction samples shows that the pseudotyped viral particles transduce with similar efficiency, regardless of the siRNA template they harbor.

Product Information

GeneNet™ Human Druggable Genome siRNA Library, Puromycin selection (Pre-Packaged)

FIV-based: Cat. # SI202B-1 (2-3 transductions)

GeneNet™ Human Druggable Genome siRNA Library, Puromycin selection (Plasmid)

FIV-based: Cat. # SI202PB-1 (50 µg)

Product Components (for pre-packaged libraries)
500-2000 µl GeneNet™ Human Druggable Genome siRNA Library, pre-packaged in VSV-G particles (5x106 ifu)
50 µl cDNA Synthesis GNF Primer (10 µM)
50 µl Fwd GNF (Forward) PCR Primer (10 µM)
50 µl Rev GNF (Reverse) PCR Primer (10 µM)
200 µl NFwd-Bio (Nested Forward Biotinylated) PCR Primer (10 µM)
200 µl NRev GNF (Nested Reverse) PCR Primer (10 µM)
200 µl NRev GNF1 (Nested Reverse 1) PCR Primer (10 µM)
200 µl NRev GNF2 (Nested Reverse 2) PCR Primer (10 µM)
200 µl NRev GNF3 (Nested Reverse 3) PCR Primer (10 µM)
20 µl Positive Control DNA
100-500 µl Packaged Positive Transduction Control (>2x105 ifu)
1 disc CD with gene/siRNA list, and data analysis program compatible with Affymetrix data file