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PMID: 6344089 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

cDNA clone for the alpha subunit of the acetylcholine receptor from the mouse muscle cell line BC3H-1.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 80 ·No. 12 ·1983-06-00 ·Pages 3845-9

Merlie JP, Sebbane R, Gardner S, Lindstrom J

Abstract

Sequences from a gene coding for mouse acetylcholine receptor alpha subunit have been inserted into a recombinant plasmid and cloned in Escherichia coli. mRNAs for acetylcholine receptors occur in low abundance in vertebrate muscle. To clone the mouse alpha-subunit cDNA, we made use of (i) a cell line, BC3H-1, that overproduces the alpha-subunit mRNA and (ii) a polysome fractionation procedure that results in enrichment of alpha-subunit mRNA. Polyadenylylated RNA was used to construct a cDNA library of 750 recombinant clones. Acetylcholine receptor-specific sequences were identified by hybrid-selected translation, followed by monoclonal antibody precipitation and peptide mapping of the translation product. One clone (pA59) that fit these criteria was found in the first 80 isolates. It had a 700-base-pair insert that was excised with Pst I. Blot-hybridization experiments with nick-translated pA59 DNA showed that BC3H-1 cells contain 100-1,000 times more alpha-subunit mRNA than does newborn or adult mouse muscle. Blot hybridization of restriction digests of mouse liver DNA revealed that pA59 is homologous to a very small number (probably one) of genomic sequences.

MeSH Terms
Animals Cell Line Cloning, Molecular DNA/metabolism DNA, Recombinant/metabolism Escherichia coli/genetics Genes Liver/metabolism Macromolecular Substances Mice Muscles/metabolism Nucleic Acid Hybridization Protein Biosynthesis RNA, Messenger/genetics Receptors, Cholinergic/genetics
Chemicals
DNA, Recombinant Macromolecular Substances RNA, Messenger Receptors, Cholinergic DNA
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Merlie J P
Sebbane R
Gardner S
Lindstrom J
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39 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.80.12.3845
Published
1983-06-00
Pages
3845-9
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC394149
Subset
IM
External Links
PubMed source DOI: 10.1073/pnas.80.12.3845 PMC Full Text View journal details
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