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Science 15 May 1992:
Vol. 256. no. 5059, pp. 992 - 997
DOI: 10.1126/science.1589782
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Articles

Science, Vol 256, Issue 5059, 992-997
Copyright © 1992 by American Association for the Advancement of Science

articles

Site-specific modification of pre-mRNA: the 2'-hydroxyl groups at the splice sites

MJ Moore and PA Sharp

Center for Cancer Research, Massachusetts Institute of Technology, Cambridge 02139.

A simple and efficient method for synthesizing long, site-specifically modified RNA molecules was developed whereby segments of RNA were joined with the use of bacteriophage T4 DNA ligase. A single hydrogen or O-methyl group was substituted for the 2'-hydroxyl group at either splice site of a nuclear pre-messenger RNA substrate. Splicing of the modified pre-messenger RNA's in vitro revealed that, although a 2'-hydroxyl is not absolutely required at either splice site, the 2'-hydroxyl at the 3' splice site is important for the second step of splicing. These results are compared to previous studies of analogous 2'-hydroxyl groups in the self-splicing Tetrahymena group I intron.


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