| Research Abstract |
1. Sex determination of DNA samples which extracted from human male and female has been performed by dot hybridization method with the use of nonradioactive Y-specific prove (pHY10) and by PCR method with the use of Y-specific primers (ASF, alphoid satellite family) and X-specific primers. The Y-specific hybridization band of the male DNA was observed in its nitrocellulose photograph pattern, which was examined by gel in situ hybridization using a nonradioactive Y-specific probe. The female showed no Y-specific hybridization band. The 172bp amplification product was detected in male DNA after PCR using Y-specific primers, but not in females. The minimum amount of blood for sex determination by both methods was 0.05mu1.
2. Dot hybridization and PCR methods can identify the sex of blood stains aged for 2 years. The detection limit with dot hybridization method and PCR method was 1mu1 and 0.05mu1, respectively.
3. It was possible to determine the sex from 10mu1 saliva (no DNA extraction) by PCR using two kinds of Y specific primers (ASF, DYZ1).
4. The effect of fixatives on sex determination by means of DNA analysis using paraffin embedded tissue fixed with five fixatives (10% formalin, buffer formalin, Carnoy's solution, 50% acetic acid, Bouin's solution) were examined. The 172bp amplification product was detected in male DNA from, except for Bouin's solution, four fixatives fixed paraffin embedded tissue with PCR Y-specific primers, but not in females. We recommend Carnoy's solution as fixative for the purpose of sex determination using paraffin embedded tissue.
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