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Comparison of three different techniques of human sperm DNA isolation for methylation assay / 华中科技大学学报(医学)(英德文版)
Journal of Huazhong University of Science and Technology (Medical Sciences) ; (6): 938-942, 2015.
Artigo em Inglês | WPRIM | ID: wpr-250316
ABSTRACT
Human sperm DNA is an important genetic and epigenetic material, whose chromatin structure differs from that of somatic cells. As such, conventional methods for DNA extraction of somatic cells may not be suitable for obtaining sperm DNA. In this study, we evaluated and compared three sperm DNA extraction techniques, namely, modified guanidinium thiocyanate method (method A), traditional phenol-chloroform method (method B), and TianGen kit method (method C). Spectrophotometry and agarose gel electrophoresis analyses showed that method A produced DNA with higher quantity and purity than those of methods B and C (P<0.01). PCR results revealed that method A was more reliable in amplifying DEAD-box polypeptide 4 (DDX4) and copy number variations (CNVs) than methods B and C, which generated false-positive errors. The results of sperm DNA methylation assay further indicated that methods A and B were effective, and the former yielded higher quantitative accuracy. In conclusion, the modified guanidinium thiocyanate method provided high quality and reliable results and could be an optimal technique for extracting sperm DNA for methylation assay.
Assuntos

Texto completo: DisponíveL Índice: WPRIM (Pacífico Ocidental) Assunto principal: Espermatozoides / DNA / Metilação de DNA / Eletroforese em Gel de Ágar / Metabolismo Limite: Adulto / Humanos / Masculino Idioma: Inglês Revista: Journal of Huazhong University of Science and Technology (Medical Sciences) Ano de publicação: 2015 Tipo de documento: Artigo

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Texto completo: DisponíveL Índice: WPRIM (Pacífico Ocidental) Assunto principal: Espermatozoides / DNA / Metilação de DNA / Eletroforese em Gel de Ágar / Metabolismo Limite: Adulto / Humanos / Masculino Idioma: Inglês Revista: Journal of Huazhong University of Science and Technology (Medical Sciences) Ano de publicação: 2015 Tipo de documento: Artigo