Yeast cytochrome b2 gene: isolation with antibody probes |
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Authors: | B Guiard J M Buhler |
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Affiliation: | 1. Centre de Génétique Moléculaire, C.N.R.S., F-91190 Gif-sur-Yvette, France;2. Laboratoire associéàl''UniversitéPierre et Marie Curie, France;3. Service de Biochimie, C.E.N. Saclay F-91191 Gif-sur-Yvette, France |
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Abstract: | An efficient technique was used to clone the gene for yeast cytochrome b2, (a nuclear encoded mitochondrial protein) using the expression vector, lambda gt11 (lac 5 nin 5 c1857 S100). This enables the insertion of yeast DNA into the beta-galactosidase structural gene (lacZ) and promotes synthesis of hybrid proteins. Screening of antigen producing clones in the lambda gt11 recombinant genomic library was achieved using antiserum against cytochrome b2 according to Young and Davis (1983) Two recombinants containing part of the gene coding for cytochrome b2 were isolated and characterized as follows: by their expression in Escherichia coli cells, examined by immuno-blotting with antibodies to pure cytochrome b2. by DNA sequence analysis. One recombinant carries a 3 Kb yeast DNA insert which contains the whole nucleotide sequence encoding cytochrome b2 and a few amino acids of the amino terminal presequence. |
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Keywords: | imported mitochondrial protein lambda gt11 vector hybrid protein expression transport des protéines mitochondriales vecteur lambda gt11 expression de protéines hybrides |
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