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Gene. 1982 Dec;20(2):255-65.

Preparation of product-specific antisera by gene fusion: antibodies specific for the product of the yeast cell-division-cycle gene CDC28.

Abstract

Antisera with specificity for the product of a yeast cell-division-cycle (CDC) gene were prepared by immunizing rabbits to a novel hybrid polypeptide. A segment of the yeast gene CDC28 was fused to the Escherichia coli lacZ gene, which encodes beta-galactosidase, by insertion of yeast sequences into the plasmid pBGF1. pBGF1 contains the lac promoter-operator and most of the lacZ gene. An EcoRI site, 16 codons upstream from the carboxyterminus of the beta-galactosidase coding region, served as a convenient splicing site for the heterologous sequences. To insure that an open reading frame be maintained between the two gene segments for some portion of the fusions, the CDC28-encoding segments were first subjected to limited digestion with nuclease BAL31 to produce random junction points. A hybrid polypeptide encoded by such a continuous open reading frame was purified from E. coli by preparative SDS-polyacrylamide gel electrophoresis and used to immunize rabbits. The resulting antisera were shown to have specificity for CDC28 gene product synthesized by cell-free translation of yeast mRNA.

PMID:
6299892
DOI:
10.1016/0378-1119(82)90044-0
[Indexed for MEDLINE]

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