| 摘要 |
The invention provides a process for preparing capped mRNAs from an RNA mixture, e.g. whole RNA isolated from a cell or tissue extract, that include s combining in a reaction mixture RNA comprising capped mRNA with a separable affinity matrix having high-affinity eIF4E bound thereto, under conditions sufficient for binding to occur between the high-affinity eIF4E and the capp ed mRNA, whereby capped mRNA is bound to the affinity matrix, separating the affinity matrix from the reaction mixture, then separating the capped mRNA from the affinity matrix. High affinity eIF4E mutants previously described a re employed in the process as well as a novel mutant disclosed and claimed herein. The mRNA preparation process is based on isolation of 5'-capped mRNA . The mRNA molecules thus isolated have intact sequences encoding the NH2- terminal ends of the proteins they encode, unlike those isolated by prior methods. In addition, use of the method isolates mRNA sequences not isolatab le by prior methods that relied on binding to polyadenylated 3'-end sequences.< /SDOAB>
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