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Title: Analysis of in vivo somatic mutations in normal human cells

We have used the APRT locus located at 16q24.3 to study the nature of loss of heterozygosity (LOH) in human T lymphocytes in vivo. T lymphocytes were isolated from blood from APRT (+/{minus}) obligated heterozygotes with known germline mutations. The cells were immediatley placed in culture medium containing 100 {mu}M 2,6-diaminopurine (DAP) to select for drug-resistant clones ({minus}/{minus}) already present. These clones were first examined using polymorphic CA microsatellite repeat markers D16S303 and D16S305 that are distal and proximal to APRT, respectively. The retention of heterozygosity of these markers is suggestive of minor changes in the APRT gene, the exact nature of which were determined by DNA sequencing. Nineteen out of 70 DAP-resistant clones from one heterozygote showed APRT sequence changes. The loss of heterozygosity of markers D16S303 and D16S305 in the remaining clones suggests LOH involving multilocus chromosomal events. These clones were then sequentially typed using additional CA repeat markers proximal and distal to APRT. The extent of LOH in these clones was found to vary from <5 cM to almost the entire 16q arm. Preliminary results suggest that there are multiple sites along the chromosome from which LOH proceeds distally in these clones. Cytogenetic analysis of 10 clonesmore » suggested mitotic recombination in 9 and deletion in one. Studies are in progress to further characterize the molecular mechanisms of LOH.« less
Authors:
; ;  [1]
  1. Indiana Univ. School of Medicine, Indianapolis, IN (United States) [and others
Publication Date:
OSTI Identifier:
134024
Report Number(s):
CONF-941009--
Journal ID: AJHGAG; ISSN 0002-9297; CNN: Grant DK38185; TRN: 95:005313-0760
Resource Type:
Journal Article
Resource Relation:
Journal Name: American Journal of Human Genetics; Journal Volume: 55; Journal Issue: Suppl.3; Conference: 44. annual meeting of the American Society of Human Genetics, Montreal (Canada), 18-22 Oct 1994; Other Information: PBD: Sep 1994
Country of Publication:
United States
Language:
English
Subject:
55 BIOLOGY AND MEDICINE, BASIC STUDIES; HUMAN CHROMOSOME 16; GENETIC MAPPING; GENES; GENE MUTATIONS; DNA SEQUENCING; GENE RECOMBINATION; STRUCTURE-ACTIVITY RELATIONSHIPS; LYMPHOCYTES; SOMATIC MUTATIONS; CLONING; MITOSIS; BIOLOGICAL MARKERS; STATISTICS