Efficient DNA Extraction from Nail Clippings Using the Protease Solution from Cucumis melo
- 733 Downloads
Owing to the increasing importance of genomic information, obtaining genomic DNA easily from biological specimens has become more and more important. This article proposes an efficient method for obtaining genomic DNA from nail clippings. Nail clippings can be easily obtained, are thermostable and easy to transport, and have low infectivity. The drawback of their use, however, has been the difficulty of extracting genomic material from them. We have overcome this obstacle using the protease solution obtained from Cucumis melo. The keratinolytic activity of the protease solution was 1.78-fold higher than that of proteinase K, which is commonly used to degrade keratin. With the protease solution, three times more DNA was extracted than when proteinase K was used. In order to verify the integrity of the extracted DNA, genotype analysis on 170 subjects was performed by both PCR–RFLP and Real Time PCR. The results of the genotyping showed that the extracted DNA was suitable for genotyping analysis. In conclusion, we have developed an efficient extraction method for using nail clippings as a genome source and a research tool in molecular epidemiology, medical diagnostics, and forensic science.
KeywordsNail clippings Polymorphism DNA extraction method Cucumis melo Keratin degradation
This study has been supported in part by a Grant-in-Aid for Young Scientists (B) from the Ministry of Education, Culture, Sports, Science and Technology of Japan (No. 19700606).
- 6.Cline, R. E., Laurent, N. M., & Foran, D. R. (2003). The fingernails of Mary Sullivan: Developing reliable methods for selectively isolating endogenous and exogenous DNA from evidence. Journal of Forensic Sciences, 48, 328–333.Google Scholar
- 7.Cervantes, J. (2004). HLA class II allele typing using DNA obtained from human fingernail clipping material. Revista de Investigacion Clinica, 56, 341–344.Google Scholar
- 9.Tahir, M. A., & Watson, N. (1995). Typing of DNA HLA-DQ alpha alleles extracted from human nail material using polymerase chain reaction. Journal of Forensic Sciences, 40, 634–636.Google Scholar
- 11.Ganapathy, C. V., & Sastri, B. N. (1939). The nature of papain. Biochemical Journal, 33, 1175–1179.Google Scholar
- 16.Yamagata, H., Masuzawa, T., Nagaoka, Y., Ohnishi, T., & Iwasaki, T. (1994). Cucumisin, a serine protease from melon fruits, shares structural homology with subtilisin and is generated from a large precursor. Journal of Biological Chemistry, 269, 32725–32731.Google Scholar
- 17.Uchikoba, T., Yonezawa, H., & Kaneda, M. (1995). Cleavage specificity of cucumisin, a plant serine protease. Journal of Biochemistry, 117, 1126–1130.Google Scholar
- 18.Kaneda, M., Yonezawa, H., & Uchikoba, T. (1995). Improved isolation, stability and substrate specificity of cucumisin, a plant serine endopeptidase. Biotechnology and Applied Biochemistry, 22, 215–222.Google Scholar
- 19.Kaneda, M., & Tominaga, N. (1975). Isolation and characterization of a proteinase from the sarcocarp of melon fruit. Journal of Biochemistry, 78, 1287–1296.Google Scholar