In vitro deletional mutagenesis for bacterial production of the 20,000-dalton form of human pituitary growth hormone

John Adelman, J. S. Hayflick, M. Vasser, P. H. Seeburg

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

The 20,000-dalton (20K) variant form of human growth hormone (hGH) present in extracts from pituitary glands differs from the major form of hGH (22K, 191 amino acids) by the deletion of amino acid residues 32-46. Using oligonucleotide-mediated mutagenesis, the DNA coding for these amino acids was deleted from the gene previously constructed by us for microbial hGH production. The DNA to be deleted was looped out by the annealing of a synthetic oligodeoxyribonucleotide to the coding strand of the hGH gene contained on recombinant phage M13 mp8 DNA. Resulting heteroduplex structures were stabilized using primer-directed in vitro DNa synthesis in the presence of T4 DNA ligase. On transformation of Escherichia coli, these hetroduplex DNAs yielded phage whose genomes contained either the original or the partially deleted hGH gene, and genotypes were distinguished by in situ plaque hybridization with synthetic oligonucleotide probes. A gene with the correct deletion was used to express the short hGH variant in E.coli.

Original languageEnglish (US)
Pages (from-to)183-193
Number of pages11
JournalDNA
Volume2
Issue number3
StatePublished - 1983
Externally publishedYes

Fingerprint

Mutagenesis
Human Growth Hormone
Growth Hormone
Genes
Bacteriophages
DNA
Amino Acids
Escherichia coli
Bacteriophage M13
DNA Ligases
Oligonucleotide Probes
Oligodeoxyribonucleotides
Pituitary Gland
Oligonucleotides
In Situ Hybridization
Genotype
In Vitro Techniques
Genome
Annealing

ASJC Scopus subject areas

  • Biochemistry
  • Genetics
  • Molecular Biology

Cite this

Adelman, J., Hayflick, J. S., Vasser, M., & Seeburg, P. H. (1983). In vitro deletional mutagenesis for bacterial production of the 20,000-dalton form of human pituitary growth hormone. DNA, 2(3), 183-193.

In vitro deletional mutagenesis for bacterial production of the 20,000-dalton form of human pituitary growth hormone. / Adelman, John; Hayflick, J. S.; Vasser, M.; Seeburg, P. H.

In: DNA, Vol. 2, No. 3, 1983, p. 183-193.

Research output: Contribution to journalArticle

Adelman, J, Hayflick, JS, Vasser, M & Seeburg, PH 1983, 'In vitro deletional mutagenesis for bacterial production of the 20,000-dalton form of human pituitary growth hormone', DNA, vol. 2, no. 3, pp. 183-193.
Adelman, John ; Hayflick, J. S. ; Vasser, M. ; Seeburg, P. H. / In vitro deletional mutagenesis for bacterial production of the 20,000-dalton form of human pituitary growth hormone. In: DNA. 1983 ; Vol. 2, No. 3. pp. 183-193.
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