Efficient 5' cap-dependent RNA purification : use in identifying and studying subsets of RNA.

摘要: Microarray-based screening technologies have revealed a larger than expected diversity of gene expression profiles for many cells, tissues, and organisms. The complexity RNA species, defined by their molecular structure, represents major new development in biology. not only carries genetic information the form templates components translational machinery protein synthesis but also directly regulates as exemplified micro-RNAs (miRNAs). Recent evidence has demonstrated that 5' capped 3' polyadenylated ends are restricted to mRNAs, they present precursors both miRNAs some antisense transcripts. In addition, 40% transcribed RNAs may lack poly(A) ends. concert with presence cap (m7 GpppN), length end plays critical role determining efficiency, stability, cellular distribution specific mRNA. short or lacking ends, escape isolation amplification oligo(dT)-based methods, provide challenge biology studies. To circumvent limitations poly(A)-dependent we developed an efficient purification system binds high-affinity variant cap-binding eIF4E. This can be used differential selection approaches isolate subsets RNAs, including those likely targets post-transcriptional regulation expression. using rapid polymerase chain reaction (PCR)- based approach.