当前位置: X-MOL 学术Clin. Chem. › 论文详情
Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)
Evaluation of Sperm DNA Integrity by Mean Number of Sperm DNA Breaks Rather Than Sperm DNA Fragmentation Index
Clinical Chemistry ( IF 9.3 ) Pub Date : 2021-12-30 , DOI: 10.1093/clinchem/hvab280
Bei Yan 1, 2 , Weicong Ye 1 , Juan Wang 3 , Shaotong Jia 2 , Xiuli Gu 4 , Hao Hu 1 , Wenpei Xiang 1 , Tongbo Wu 1 , Xianjin Xiao 1
Affiliation  

Background Sperm DNA integrity is crucial for normal fertilization, implantation, and embryo development. Several assays are available to assess sperm DNA fragmentation but are limited by high price, complicated processes, and low accuracy. Methods We developed a secondary amplification detection system based on terminal deoxynucleotidyl transferase and endonuclease IV, which could efficiently measure the number of 3'-OH (equivalent to the number of breakpoints). We applied this detection system in single stranded DNA with standard concentrations to obtain the standard curve. We then broke the double stranded genomic DNA by ultrasound and enzyme digestion and used the detection system to monitor the increase of DNA breakpoints. Finally, we used this method to measure the mean number of sperm DNA breakpoints (MDB) in 80 sperm samples. Results We successfully measured the number of 3'-OH in single stranded DNA with standard concentration and obtained the standard curve. The linear range for the number of DNA breakpoints was from 0.1 nM to 15 nM. The detection method was successfully validated on λ DNA and 80 human sperm samples. The results of real clinical samples revealed that the mean number of DNA breakpoints (MDB) had a stronger relevance with the sperm motility and clinical pregnancy outcomes than the commonly used parameter of DNA fragmentation index (DFI). Conclusion We have developed a straight-forward method for direct measurement of the mean number of DNA breakpoints in sperms. The method has advantages of short time-consumption, simple operation, high analytical sensitivity, and low requirement for instrumentation, which makes it conducive to clinical application. The proposed new parameter (MDB) could be a more direct, accurate and clinically significant indicator for evaluating the sperm DNA integrity.

中文翻译:

通过平均精子 DNA 断裂数而不是精子 DNA 片段化指数评估精子 DNA 完整性

背景 精子 DNA 的完整性对于正常的受精、植入和胚胎发育至关重要。有几种检测方法可用于评估精子 DNA 片段化,但受到价格高、过程复杂和准确性低的限制。方法我们开发了一种基于末端脱氧核苷酸转移酶和核酸内切酶IV的二级扩增检测系统,可以有效地检测3'-OH的数量(相当于断点的数量)。我们将该检测系统应用于具有标准浓度的单链 DNA 以获得标准曲线。然后我们通过超声波和酶消化破坏双链基因组DNA,并使用检测系统监测DNA断点的增加。最后,我们使用这种方法测量了 80 个精子样本中精子 DNA 断点 (MDB) 的平均数。结果我们成功地用标准浓度测定了单链DNA中3'-OH的数目,得到了标准曲线。DNA 断点数量的线性范围为 0.1 nM 至 15 nM。该检测方法在 λ DNA 和 80 个人类精子样本上成功验证。实际临床样本的结果表明,与常用的DNA断裂指数(DFI)参数相比,DNA断裂点的平均数(MDB)与精子活力和临床妊娠结局的相关性更强。结论 我们开发了一种直接测量精子中 DNA 断裂点平均数的直接方法。该方法具有耗时短、操作简单、分析灵敏度高、对仪器要求低等优点,有利于临床应用。提出的新参数 (MDB) 可能是评估精子 DNA 完整性的更直接、准确和具有临床意义的指标。
更新日期:2021-12-30
down
wechat
bug