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3'-end labeling of RNA with recombinant yeast poly(A) polymerase
JournalArticle (Originalarbeit in einer wissenschaftlichen Zeitschrift)
 
ID 153584
Author(s) Lingner, J.; Keller, W.
Author(s) at UniBasel Keller, Walter
Year 1993
Title 3'-end labeling of RNA with recombinant yeast poly(A) polymerase
Journal Nucleic Acids Research
Volume 21
Number 12
Pages / Article-Number 2917-20
Keywords Base Sequence; Deoxyadenine Nucleotides/*metabolism; Hela Cells/chemistry; Humans; Kinetics; Molecular Sequence Data; Polynucleotide Adenylyltransferase/*metabolism; RNA/isolation & purification/*metabolism; RNA Ligase (ATP)/metabolism; RNA; Small Nuclear/metabolism; Recombinant Proteins; Saccharomyces cerevisiae/*enzymology; Substrate Specificity
Abstract Two commonly used methods to end-label RNA-molecules are 5'-end labeling by polynucleotide kinase and 3'-end labeling with pCp and T4 RNA ligase. We show here that RNA 3'-ends can also be labeled with the chain-terminating analogue cordycepin 5'-triphosphate (3'-deoxy-ATP) which is added by poly(A) polymerase. For a synthetic RNA it is shown that 40% of cordycepin becomes incorporated when the nucleotide is used at limiting concentrations and that with an excess of cordycepin 5'-triphosphate essentially all the RNA becomes modified at its 3'-end. The reaction is complete within minutes and the RNA product is uniform and suitable for sequence analysis. The efficiency of labeling varies with different RNA-molecules and is different from RNA ligase. Poly(A) polymerase preferentially labels longer RNA-molecules whereas short RNA-molecules are labeled more efficiently by T4 RNA ligase.
Publisher Oxford University Press
ISSN/ISBN 0305-1048 ; 1362-4962
edoc-URL http://edoc.unibas.ch/dok/A5257982
Full Text on edoc Available
Digital Object Identifier DOI 10.1093/nar/21.12.2917
PubMed ID http://www.ncbi.nlm.nih.gov/pubmed/7687347
Document type (ISI) Journal Article
 
   

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