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Comparison of three different techniques of human sperm DNA isolation for methylation assay

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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.

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Authors and Affiliations

  1. Center of Reproductive Medicine, Institute of Family Planning Research, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China

    Hong-fang Yuan  (袁红方), Martin Kuete  (马 丁), Fan Yang  (杨 帆), Zhi-yong Hu  (扈智勇), Bo-zhen Tian  (田浡臻), Hui-ping Zhang  (章慧平) & Kai Zhao  (赵 凯)

  2. Family Planning Women and Children Service Center of ZengDu District, Suizhou, 441300, China

    Li Su  (苏 莉)

Authors
  1. Hong-fang Yuan  (袁红方)
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  2. Martin Kuete  (马 丁)
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  3. Li Su  (苏 莉)
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  4. Fan Yang  (杨 帆)
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  5. Zhi-yong Hu  (扈智勇)
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  6. Bo-zhen Tian  (田浡臻)
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  7. Hui-ping Zhang  (章慧平)
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  8. Kai Zhao  (赵 凯)
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Corresponding author

Correspondence to Kai Zhao  (赵 凯).

Additional information

Both authors contributed equally to this work.

This project was supported by the National Natural Science Foundation of China (No. 81370755).

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Yuan, Hf., Kuete, M., Su, L. et al. Comparison of three different techniques of human sperm DNA isolation for methylation assay. J. Huazhong Univ. Sci. Technol. [Med. Sci.] 35, 938–942 (2015). https://doi.org/10.1007/s11596-015-1532-0

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  • DOI: https://doi.org/10.1007/s11596-015-1532-0

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