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Nucleic Acids Research, 1982, Vol. 10, No. 15 4467-4482
© 1982

MOLECULAR BIOLOGY

Chemical synthesis and cloning of a gene for human ß-urogastrone

John Smith, E. Cook, Ian Fotheringham, Sally Pheby, Roger Derbyshire, Michael A.W. Eaton1, Mike Doel1, David M.J. Lilley2, John F. Pardon, Thakor Patel1, Hilary Lewis3 and Leslie D. Bell

Searle Research and Development Lane End Road, High Wycombe, Bucks HP12 4HL, UK

Received June 14, 1982. Accepted July 6, 1982.

A DNA duplex coding for the 53 amino acids of human ß-urogastrone has been synthesised. Computer assisted design of the gene included restriction endonuclease sites for plasmid insertion, a termination codon and two triplets coding for lysine at the 5'-end of the structural gene.

The synthesis involved preparation of 23 oligodeoxyribo-nucleotides by phosphotriester procedures coupled to rapid HPLC techniques. The gene was constructed in two halves by enzymatic ligation of the oligonucleotides and cloned into a specially constructed chimeric plasmid vector. Escherichia coli K12 MRC8 was transformed by the plasmid and clones containing the full gene sequence were isolated and characterised.

1Present address: Celltech, 250, Bath Road, Slough, Bucks, UK

2Present address: Biochemistry Department, Medical Sciences Institute, University of Dundee, Dundee DD1 4HN, UK

3Present address: Department of Microbiology, University of Sheffield, Sheffield, UK


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