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Isolation of mouse N‐CAM‐related cDNA: detection and cloning using monoclonal antibodies.
Author(s) -
Goridis C.,
Hirn M.,
Santoni M.J.,
Gennarini G.,
DeagostiniBazin H.,
Jordan B.R.,
Kiefer M.,
Steinmetz M.
Publication year - 1985
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1985.tb03676.x
Subject(s) - cloning (programming) , biology , computer science , programming language
Clones coding for the mouse neural cell adhesion molecule (N‐CAM) were isolated from a cDNA library prepared in the expression vector lambda gt 11 from mRNA extracted from a mouse neuroblastoma cell line. This library was screened with two anti‐N‐CAM monoclonal antibodies directed against different sites on the molecule and with rabbit anti‐N‐CAM serum. Two clones were identified with the first monoclonal antibody, three with the second one, none reacted with both. The relevance of these cDNA clones to N‐CAM was confirmed by several observations. First, cDNA sequences detected with one monoclonal antibody cross‐hybridized with those identified by the other antibody. Second, the different fusion proteins all bound the rabbit serum in addition to one monoclonal antibody. Finally, the probes hybridized to discrete mRNA species of sufficient lengths to code for the very large N‐CAM polypeptides in RNA preparations from N‐CAM‐expressing, but not from N‐CAM‐negative cells. An additional mRNA species not seen in embryonic brain was expressed in adult mouse brain. Genomic blot experiments indicated that sequences corresponding to one of our probes are present only a few times in the mouse genome.