DNA sequence analysis of hprt mutants persisting in peripheral blood of cynomolgus monkeys more than two years after ENU treatment
✍ Scribed by P.R. Harbach; S.S. Mattano; D.M. Zimmer; A.L. Filipunas; Y. Wang; C.S. Aaron
- Publisher
- John Wiley and Sons
- Year
- 1999
- Tongue
- English
- Weight
- 62 KB
- Volume
- 33
- Category
- Article
- ISSN
- 0893-6692
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✦ Synopsis
We have been studying in vivo mutagenesis at the hypoxanthine phosphoribosyl transferase (hprt) locus in cynomolgus monkey T-lymphocytes. This primate model allows us to study mutations and their kinetics under well-controlled conditions. Previously, we reported mutations detected at various times after intraperitoneal treatment with ethylnitrosourea (ENU, 77 mg/kg). At 832 days after that first treatment, the monkey received a second dose of 77 mg/kg ENU. Up to 1,331 days after the second treatment, the T-cell mutant frequency (44.2 ϫ 10 Ϫ6 ) was still 26-fold higher than background (1.7 ϫ 10 Ϫ6 ), suggesting that mutants persisted in the peripheral blood. Mutant clones from Days 974, 1,164, and 1,311 after the second treatment were selected in thioguanine. Hprt cDNA was prepared from a cell lysate, PCR-amplified, and sequenced. Of 45 mutants, 30 yielded PCR product and 26 were sequenced. Base substitutions were found in 21 (81%) of the 26 mutants