August’s issue of CSH Protocols is now available, and one of the featured protocols this month comes from Inder Verma’s lab, and covers the Design and Cloning of an shRNA into a Lentiviral Vector. Combining the specificity of small interfering RNA (siRNA) silencing with the versatility of lentiviral vectors gives researchers a powerful tool for the investigation of gene function both in vivo and in vitro. There’s also an alternative method available. In the featured method, one undesirable consequence of this procedure is that the siRNA target sequence is also present in the mRNA expressing the marker gene, resulting in somewhat lower expression of the marker. In the alternative method, the position of the silencing cassette is upstream of the marker expression cassette, thus avoiding down-regulation of the marker. But, because the silencing cassette is not in the 3′ LTR, only one copy of the silencing cassette is delivered per viral particle (as opposed to two copies in the featured method).

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