Description
Quantity: 1 kit
Unit Size: Each
Features
• Measure the poly(A) tail of multiple RNAs in under 3 hrs using common lab equipment
• Accurate measurement of poly(A) tail length compared to other assays
• One kit includes all necessary reagents for 5 G/I tailing, 20 RT, and 80 PCR reactionsTechnologyThe Poly(A) Tail-Length Assay Kit uses four key steps to enable poly(A) tail-length determination (see figure).
• Step 1—poly(A) polymerase adds a limited number of guanosine and inosine residues to the 3'-ends of poly(A)-containing RNAs.
• Step 2—the tailed-RNAs are converted to DNA through reverse transcription using the newly added G/I tails as the priming sites.
• Step 3—PCR amplification products are generated using two primer sets. A gene-specific forward and reverse primer set designed upstream of the polyadenylation site (e.g., the 3'-UTR) is produced as a control for the gene-of-interest. The second set of primers uses the gene-specific forward primer and the universal reverse primer provided with the kit to generate a product that includes the poly(A) tails of the gene-of-interest.
• Step 4—the PCR products are separated on an agarose or polyacrylamide gel (see figure).Selected references
• Parker, R. and Song, H. (2004)Nat Struct Mol Biol.11 121-127.
• Brodersen, D.E.et al.(2008)Biochim Biophys Acta.1779, 532-537.
• Wu, L., Fan J. and Belasco, J.G. (2006)Proc Natl Acad Sci U S A. 103, 4034-4039.
• Izaurralde, E.et al.(2009)RNA15, 21-32.
• Martin, G. and Keller, W. (1998)RNA4, 226-230.
• Gauss-Müller, V.et al.(2001)Nucleic Acids Res.29 E57-7
