An improved rapid enzymatic method of RNA sequencing using chemical modification.

Autor: Mazo AM, Mashkova TD, Avdonina TA, Ambartsumyan NS, Kisselev LL
Jazyk: angličtina
Zdroj: Nucleic acids research [Nucleic Acids Res] 1979 Dec 20; Vol. 7 (8), pp. 2469-82.
DOI: 10.1093/nar/7.8.2469
Abstrakt: A version of rapid gel sequencing procedure based on the analysis of partial endonuclease hydrolizates of chemically modified 5'-32P-labelled RNA is suggested. Complete and selective modification of cytidilic residues by a methoxyamine-bisulfite mixture leads to the unfolding of the RNA secondary structure and, due to this effect, to the generation of a more uniform set of fragments after partial RNAase hydrolysis. The position of cytidines in an RNA sequence can be determined by restricting the hydrolysis of phosphodiester bonds between the modified CMP residues and their 3'-neighbours with T2 and A RNAases. The method was verified with tRNATrp (yeast) and 5S RNA (rat liver and yeast).
Databáze: MEDLINE