Direct detection of small RNAs using splinted ligation
作者： Patricia A Maroney , Sangpen Chamnongpol , Frédéric Souret , Timothy W Nilsen
关键词:
摘要: This protocol describes a method for direct labeling and detection of small RNAs present in total RNA by splinted ligation. The assay uses RNA-specific bridge oligonucleotide to form base pairs with the 5′-end-radiolabeled ligation oligonucleotide. captured is directly labeled Detection performed denaturing gel electrophoresis autoradiography or phosphorimaging. has been successfully used study expression various classes biological from nanogram microgram amounts without an amplification step. It significantly simpler perform more sensitive than either northern blotting ribonuclease protection assays. Once oligonucleotides have synthesized extracted, procedure can be completed 6 h.
nature.com
sci-hub.se 参考文章(11)
Melissa J. Moore, Charles C. Query, Joining of RNAs by splinted ligation. Methods in Enzymology. ,vol. 317, pp. 109- 123 ,(2000) , 10.1016/S0076-6879(00)17009-0
David P Bartel, MicroRNAs: Genomics, Biogenesis, Mechanism, and Function Cell. ,vol. 116, pp. 281- 297 ,(2004) , 10.1016/S0092-8674(04)00045-5
Alexander Hüttenhofer, Jörg Vogel, Experimental approaches to identify non-coding RNAs Nucleic Acids Research. ,vol. 34, pp. 635- 646 ,(2006) , 10.1093/NAR/GKJ469
P. A. Maroney, S. Chamnongpol, F. Souret, T. W. Nilsen, A rapid, quantitative assay for direct detection of microRNAs and other small RNAs using splinted ligation. RNA. ,vol. 13, pp. 930- 936 ,(2007) , 10.1261/RNA.518107
R. S. PILLAI, MicroRNA function: multiple mechanisms for a tiny RNA? RNA. ,vol. 11, pp. 1753- 1761 ,(2005) , 10.1261/RNA.2248605
Patricia A Maroney, Yang Yu, Jesse Fisher, Timothy W Nilsen, Evidence that microRNAs are associated with translating messenger RNAs in human cells Nature Structural & Molecular Biology. ,vol. 13, pp. 1102- 1107 ,(2006) , 10.1038/NSMB1174
Søren Peter Jonstrup, Jørn Koch, Jørgen Kjems, A microRNA detection system based on padlock probes and rolling circle amplification RNA. ,vol. 12, pp. 1747- 1752 ,(2006) , 10.1261/RNA.110706
Jinmai Jiang, Eun Joo Lee, Yuriy Gusev, Thomas D Schmittgen, Real-time expression profiling of microRNA precursors in human cancer cell lines Nucleic Acids Research. ,vol. 33, pp. 5394- 5403 ,(2005) , 10.1093/NAR/GKI863
J. SHINGARA, An optimized isolation and labeling platform for accurate microRNA expression profiling RNA. ,vol. 11, pp. 1461- 1470 ,(2005) , 10.1261/RNA.2610405
Caifu Chen, Dana A Ridzon, Adam J Broomer, Zhaohui Zhou, Danny H Lee, Julie T Nguyen, Maura Barbisin, Nan Lan Xu, Vikram R Mahuvakar, Mark R Andersen, Kai Qin Lao, Kenneth J Livak, Karl J Guegler, Real-time quantification of microRNAs by stem–loop RT–PCR Nucleic Acids Research. ,vol. 33, pp. 179- ,(2005) , 10.1093/NAR/GNI178