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Cloning and expression of a collagen-analog-encoding synthetic gene in Escherichia coli
Authors:Ina Goldberg  Anthony J Salerno  Thomas Patterson  Jon I Williams
Abstract:A family of totally synthetic genes coding for multiple tandem repeats of the amino acid sequence (Gly-Pro-Pro) has been prepared and inserted into the ClaI cloning site of the expression vector pJL6. A representative recombinant plasmid, pAC1, with an insert of about 340 bp was established in an Escherichia coli strain bearing a defective λ prophage, to study expression of the CII-collagen analog fusion protein produced from pAC1 upon heat induction. Authentic fusion protein production was demonstrated by nucleotide sequencing, Northern-blot analysis, and in vivo synthesis. Conversion of a wild-type rpoH allele to the rpoH165 mutation was shown to suppress proteolysis of the unstable fusion protein.
Keywords:Recombinant DNA  oligodeoxyribonucleotides  gene construction  plasmid vector  phage λ    promoter  represser  poly(Gly-Pro-Pro)
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