Sequence analysis of nonradioactive RNA fragments by periodate-phosphatase digestion and chemical tritium labeling: characterization of large oligonucleotides and oligonucleotides containing modified nucleosides.

Nucleic acids research ·Vol. 1 ·No. 9 ·1974-09-00 ·Pages 1121-41

Randerath K, Randerath E, Chia LS, Gupta RC, Sivarajan M

Abstract

A tritium derivative method for sequence analysis of polyribonucleotides is detailed, which is based on borotritide reduction of oligonucleotide dialdehydes generated by treatment of polyribonucleotides with alkaline phosphatase and excess periodate at pH 8 (borate buffer; no primary amine present in the reaction mixture). While neither phosphatase nor periodate possess any intrinsic exonuclease activity their combination mimics an RNA-specific exonuclease ("pseudo-exonuclease"). Procedures are described for separation and characterization of tritiumlabeled oligonucleotide derivatives. The sequence is deduced by identification of labeled 3'-termini following separation of the reduced nucleotide intermediates according to chain length. The sensitivity of the method is indicated by the fact that as little as 0.01 O.D.260 unit of a nonradioactive decanucleotide is sufficient for sequence determination.

MeSH Terms

Alkaline Phosphatase Base Sequence Biochemical Phenomena Biochemistry Chromatography, Thin Layer Methods Oligonucleotides/analysis Periodic Acid RNA Ribonucleases Time Factors Tritium

Chemicals

Oligonucleotides Tritium Periodic Acid RNA Ribonucleases Alkaline Phosphatase

Authors & Affiliations

5 authors, click to expand affiliations / ORCID

Randerath K

Randerath E

Chia L S

Gupta R C

Sivarajan M

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33 references, click to expand

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Article Info

Journal

Nucleic acids research

Abbr.

Nucleic Acids Res

ISSN

0305-1048

DOI

10.1093/nar/1.9.1121

Published

1974-09-00

Pages

1121-41

Language

English

Region

England

NLM ID

0411011

PMCID

PMC344335

Subset

External Links

PubMed source DOI: 10.1093/nar/1.9.1121 PMC Full Text View journal details

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Sequence analysis of nonradioactive RNA fragments by periodate-phosphatase digestion and chemical tritium labeling: characterization of large oligonucleotides and oligonucleotides containing modified nucleosides.

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