Molecular nature of spontaneous mutations at the hypoxanthine-guanine phosphoribosyltransferase (hprt) locus in chinese hamster ovary cells

Z. Xu, Yongjia Yu, J. L. Schwartz, M. L. Meltz, A. W. Hsie

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

The hypoxanthine-guanine phosphoribosyltransferase (hprt) locus has been widely used as a selectable genetic marker for studies of mammalian cell mutagenesis. We report here the spontaneous mutation spectrum at the hprt locus in 64 independently isolated mutants of Chinese hamster ovary (CHO) cells. All nine hprt exons were simultaneously analyzed via multiplex polymerase chain reaction (PCR) for rapid detection of gene deletions or insertions. Structural point mutations were identified by direct sequence analysis of the PCR amplified cDNA. The molecular nature of RNA splicing errors and insertions was analyzed by solid-phase direct exon sequencing. Single base substitutions were found in 24 mutants (38%), of which 21 were missense and 3 were nonsense mutations. Transversions were about twice as frequent as transitions. Fifteen mutants (23%) had deletions involving either intragenic small fragments (2), single exons (9), or multiple exons (4). The majority of deletion breakpoints (71%) were located in regions surrounding exons 4, 5, and 6. RNA splicing mutations were observed in 15 mutants (23%) and effected exons 3-8; most (6/15) resulted in the loss of exon 7. Two insertion mutants, one with a 209 bp insert in exon 4 and the other with a 88 bp insert accompanied by a 24 bp deletion in exon 6, represent novel mutations reported for the first time in spontaneous mutants of the mammalian hprt gene.

Original languageEnglish (US)
Pages (from-to)127-138
Number of pages12
JournalEnvironmental and Molecular Mutagenesis
Volume26
Issue number2
DOIs
StatePublished - 1995

Fingerprint

Hypoxanthine Phosphoribosyltransferase
Cricetulus
Ovary
mutation
Exons
Cells
Mutation
polymerase chain reaction
RNA Splicing
RNA
Polymerase chain reaction
gene
genetic marker
Genes
mutant
Mutagenesis
substitution
Nonsense Codon
Insertional Mutagenesis
Multiplex Polymerase Chain Reaction

Keywords

  • CHO cells
  • hprt
  • Molecular mutagenesis
  • Multiplex PCR
  • Mutation spectrum

ASJC Scopus subject areas

  • Environmental Science(all)
  • Environmental Chemistry
  • Genetics
  • Genetics(clinical)
  • Toxicology
  • Health, Toxicology and Mutagenesis

Cite this

Molecular nature of spontaneous mutations at the hypoxanthine-guanine phosphoribosyltransferase (hprt) locus in chinese hamster ovary cells. / Xu, Z.; Yu, Yongjia; Schwartz, J. L.; Meltz, M. L.; Hsie, A. W.

In: Environmental and Molecular Mutagenesis, Vol. 26, No. 2, 1995, p. 127-138.

Research output: Contribution to journalArticle

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