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siRNA Expression Vectors with Selectable Markers

For Long Term Studies in Mammalian Cells


Six new pSilencer siRNA expression vectors are now available, each with an antibiotic resistance gene to facilitate selection in mammalian cultured cells. These new vectors are ideal for the long-term study of gene specific knockdown. Gene specific silencing in mammilian cells can be readily accomplished by inducing the RNA interference pathway with siRNAs (see Gene Specific Silencing by RNAi). Advantages of using siRNA expression vectors over siRNAs prepared by chemical synthesis, in vitro transcription or RNase III digestion of long dsRNA include the ability to perform long term gene silencing studies and eliminating the need to work directly with RNA.


How siRNA Expression Vectors Work
Vectors that express siRNAs within mammalian cells typically use an RNA polymerase III promoter to drive expression of a short hairpin RNA that mimics the structure of an siRNA (See Why RNA Polymerase III Promoters? in sidebar). The insert that encodes this hairpin is designed to have two inverted repeats separated by a short spacer sequence (Figure 1). One inverted repeat is complementary to the mRNA to which the siRNA is targeted. A string of thymidines added to the 3' end serves as a pol III transcription termination site. (See Design of Vector Encoded siRNAs in sidebar). Once inside the cell, the vector constitutively expresses the hairpin RNA, which induces silencing of the target gene.



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