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PMID: 6410388 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Cloning of human purine-nucleoside phosphorylase cDNA sequences by complementation in Escherichia coli.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 80 ·No. 14 ·1983-07-00 ·Pages 4281-5

Goddard JM, Caput D, Williams SR, Martin DW

Abstract

We have obtained cDNA clones that contain the entire coding region of the human purine-nucleoside phosphorylase (PNP; EC 2.4.2.1) mRNA. The cDNA sequences were generated by reverse transcription of PNP-enriched mRNA obtained by immunoadsorption of HeLa cell polyribosomes with monospecific antibody to human PNP. cDNA molecules that were close in length to PNP mRNA were separated by agarose gel electrophoresis and inserted into the Pst I site of the plasmid pBR322. Plasmid DNA from the pooled clones was used to transform PNP-deficient Escherichia coli cells, and those transformants that phenotypically expressed PNP were isolated on selective media. The presence of human PNP in the selected bacterial cells was detected by immunoprecipitation with human PNP antibody.

MeSH Terms
Base Sequence Cloning, Molecular DNA/analysis Escherichia coli/genetics Genes Genetic Complementation Test Humans Nucleic Acid Hybridization Pentosyltransferases/genetics Plasmids Protein Biosynthesis Purine-Nucleoside Phosphorylase/genetics,isolation & purification RNA, Messenger/genetics
Chemicals
RNA, Messenger DNA Pentosyltransferases Purine-Nucleoside Phosphorylase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Goddard J M
Caput D
Williams S R
Martin D W
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35 references, click to expand
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.80.14.4281
Published
1983-07-00
Pages
4281-5
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC384021
Subset
IM
Grants
NIAID NIH HHS · AI 17435 · United States
External Links
PubMed source DOI: 10.1073/pnas.80.14.4281 PMC Full Text View journal details
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