MICROCHEMICAL JOURNAL
·
2025
·
IF 5.1
Shihua Luo,Lifeng Zhang,Xiaohe Zhang,Jiezhen Pan,Mei Wang,Ye Zhang,Aiping Qin,Wencheng Chen
The integrity of sperm DNA is essential for proper fertilization and embryonic development. Various methods exist to evaluate sperm DNA fragmentation. However, they are frequently hindered by high costs, complex procedures, and low precision. Herein, a simple and sensitive fluorescence biosensor was developed using the terminal deoxynucleotidyl transferase (TdT) and CRISPR/Cas12a system for analyzing sperm DNA fragmentation. This biosensor efficiently measures the number of 3′-OH, which correspond to the mean number of sperm DNA breakpoints (MDB). In the presence of TdT, the dATP nucleotide polymerized at the 3′-OH end of the primer, generating numerous polyA-tails, which were detected by CRISPR-guided RNA (crRNA) at the same time. Ultimately, the trans -cleavage activity of CRISPR/Cas12a was activated, which then cleaved the spherical nucleic acids (SNA) reporter, generating amplified fluorescence signal. The proposed method exhibited a broad linear detection for the quantity of 3′-OH ends in single-stranded DNA, from 1 fM to 10 pM, with a limit of detection of 2.5 fM. Moreover, the platform was successfully validated with clinical sperm samples and demonstrated a stronger correlation with sperm motility compared to traditional DNA fragmentation index (DFI) test. Therefore, the developed method offers advantages such as ease of use, elevated sensitivity, and mimimal instrumentation requirements, making it suitable for clinical application.
研究领域:
生殖生物学
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分子诊断学
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基因编辑技术
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产品关联
DOI
高影响因子
SENSORS AND ACTUATORS A-PHYSICAL
·
2023
·
IF 4.6
Xiaoliang Zhang,Jinxian Wang,Xin Lyu,Tianhang Yang,Wei Liu,Bidou Wang,Gangyin Luo
Droplet-based microfluidic (DMF) technology is widely used in genetic analysis. Among multiple droplet generation techniques, acoustic droplet ejection (ADE) is a non-contact, high-precision method that utilizes the energy of focused ultrasound. The ejection of a single droplet of controlled size, the performance of multi-droplet ejections and the effects of the ultrasound energy on nucleic acid molecules in the droplets, are the main research interests in droplet-based genomics. In this work, a focused ultrasonic transducer with a frequency of 20 MHz was used, and we achieved droplet ejection successfully. Various experiments were carried out to study ADE and the beneficial results were obtained, including the characterization of ADE conditions, the relationship between droplet volume and parameters (including power, duration and distance), the energy required for ADE, and the effects of ultrasound energy on DNA strands, etc. It was first noticed that the superior limit of the droplet size could be increased by adjusting the height of the transducer. In addition, multi-droplet ejections at the optimal delay time were realized with good linearity. Above all, we demonstrated that the nucleic acid molecules remained their integrity when the exciting power was the maximum and the exciting duration was the longest in our system. In summary, ADE was proved to have an important role and a vast prospect in DMF technology for genomics-related applications.
研究领域:
微流控技术
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基因组学
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生物技术
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超声技术
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产品关联
DOI
高影响因子
ECOTOXICOLOGY AND ENVIRONMENTAL SAFETY
·
2019
·
IF 4.527
Cong Yuan,Lihong Wang,Long Zhu,Benhui Ran,Xue Xue,Zaizhao Wang
Ubiquitous BPA exposure resulted in DNA methylation errors and oxidative stress . Numerous studies have demonstrated that oxidative stress can lead to changes in DNA methylation levels and supplementation with antioxidants, including N-acetylcysteine (NAC), was able to restore these changes. Our previous study supposed that BPA-induced de novo synthesis of glutathione (GSH) promoted DNA methylation process in Gobiocypris rarus testes . To validate this conjecture and explore the protective effects of NAC on BPA toxicity, the present study was carried out. Adult male G. rarus was treated with 225 μg L −1 BPA and/or NAC for 7 days. The sperm motility and DNA integrity of G. rarus were determined. Meanwhile, the levels of 5-methylcytosine (5mC), GSH, hydrogen peroxide (H 2 O 2 ), DNA methyltransferase proteins (DNMTs), γ-glutamyl cysteine synthetase (GCS), S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), homocysteine (HCY), nicotinamide adenine dinucleotide phosphate (NADPH) and cysteine in the testes were detected. Furthermore, the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) were measured. Results indicated that NAC addition resulted in increase of cysteine contents and partially inhibited the BPA-induced DNA hypermethylation of G. rarus testes. In addition, the changes in DNA methylation levels in the testes after BPA and/or NAC treatment might be controlled by DNA methylation process that mediated by DNMTs. Moreover, BPA exposure caused oxidative stress in the testes and the elimination of H 2 O 2 might be mainly accomplished by CAT while it changed to mainly through GPx after NAC supplement. Finally, the positive response of testicular antioxidant enzyme system and the antioxidant activity of NAC itself protected sperm motility and DNA integrity from oxidative damage in each group.
研究领域:
环境科学
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毒理学
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分子生物学
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水生生物学
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产品关联
DOI
PubMed
高影响因子
Antioxidants
·
2025
·Vol. 14
·期 6
·630
·
IF 6.6
Qian-Qiu Liu,Xiong Li,Jia-Hao Li,Yang Zhou,Ming-Kai Lei,Wei-Qi Yin,You-She Ren,Chun-He Yang,Chun-Xiang Zhang
The concurrent occurrence of summer heat stress and the non-breeding season has significantly impaired the semen quality of rams. Currently, there exists no straightforward and efficient method to address this issue. In this study, we demonstrate that two consecutive administrations of melatonin implants significantly mitigate the adverse effects of summer heat stress and the non-reproductive season on rams. Our findings indicate that implantation of exogenous melatonin enhances semen quality by improving sperm DNA integrity, mitochondrial integrity, and decreasing the proportion of abnormal sperm, as compared to control rams. This improvement is ascribed to the alleviation of oxidative stress and the optimized regulation of endocrine hormone levels in both serum and seminal plasma. Further exploration of the regulatory mechanism reveals that melatonin can also influence the tryptophan metabolism pathway. Additionally, our study revealed that certain indices and metabolites are strongly correlated with semen quality and can potentially serve as indicators for research aimed at improving semen quality. Notably, this is the first time that differences in tryptophan metabolites between serum and seminal plasma have been elucidated. All the above information suggests that melatonin implantation can protect sperm from heat stress by optimizing the blood and semen microenvironment.
研究领域:
动物繁殖学
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内分泌学
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生殖生物学
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生物化学
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应激生理学
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DOI
PubMed
高影响因子
AQUATIC TOXICOLOGY
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2021
·
IF 4.964
Wei Shi,Shuge Sun,Yu Han,Yu Tang,Weishang Zhou,Weixia Zhang,Xueying Du,Lin Huang,Guangxu Liu
By employing external fertilization (broadcast spawning) as a mating strategy, the gametes and subsequent fertilization of various marine invertebrates are directly subjected to pollution. Although microplastics (MPs) are ubiquitous in marine environments, their potential effects on the fertilization of broadcast spawners remain largely unknown. Therefore in this study, the impacts of polystyrene MPs on the fertilization success of broadcast spawning bivalve ( Tegillarca granosa ) were investigated. In order to reveal the underlying mechanisms affecting fertilization, the sperm swimming performance, sperm ATP status, sperm viability, DNA integrity, gamete collision probability, gamete fusion efficiency, enzymatic antioxidants, and key ion transport enzyme activities were analyzed. The results showed that MPs weakened the sperm swimming performance through reducing ATP production and cell viability, thus leading to the decreased probability of gamete collision. Furthermore, MPs affected ion transport in the gametes by inducing oxidative stress, which resulted in gamete fusion failure. In conclusion, this study demonstrates that MPs could significantly decrease the fertilization success of T. granosa through reducing gamete collision and lowering gamete fusion efficiency.
研究领域:
海洋生物学
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环境科学
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生殖生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）