Production of transgenic mice expressing human ZPA and its application to sperm function diagnosis

人ZPA转基因小鼠的制备及其在精子功能诊断中的应用

基本信息

批准号：
10470350
负责人：
KOYAMA Koji
金额：
$ 7.81万
依托单位：
Hyogo college of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-10470350/
关键词：
Zona pellucida Pig ZPA Transgenic mice Male infertility Sperm function diagnosis In vitro fertilization ZPA 受精受精能獲得精子レセプター先体反応不妊症

项目摘要

The objective of this study is to produce zona pellucida for use in human sperm function diagnosis. We attempted to make transgenic mice expressing pig ZPA protein to which human sperm binds. The reason why we selected pig ZPA instead of human is that another group had already started a similar study using the human ZPA gene. The transgene was constructed with the promoter region of mouse ZPA (1.4kb) and the minigene including pig ZPA full-length DNA (6.8kb) and polyadenylation signal of rabbit β-globin (0.5kb). The promoter DNA fragment of mouse ZPA was kindly provided by Dr.J.Dean of NIH inUA.In order to introduce the transgene into mouse embryos BDF1 female mice were treated with PMS/hCG for superovulation before mating. The injected embryos was implanted in the oviducts of pseudo-pregnant mice. The incorporation of the transgene were examined by PCR analysis using DNA extracted from the tails. The transgene was detected in four newly born mice. Two (_<2, >8) of them transferred the transgene genes to their progenies. The corresponding transcripts were detected not only in ovarian tissue but also in other tissues including liver, heart, lung. But protein expression was only observed in oocytes and zona pellucida. The heterologous sperm binding capacity of the zona pellucida from the transgenic mice is currently being investigated.

这项研究的目的是生产Zona pellucida用于人类精子功能诊断。我们试图使表达人类精子结合的猪ZPA蛋白的转基因小鼠。我们选择猪ZPA而不是人类的原因是，另一组已经使用人ZPA基因开始了类似的研究。通过小鼠ZPA（1.4KB）和小型的启动子区域构建转化，包括PIG ZPA全长DNA（6.8Kb）和兔β-蛋白（0.5KB）的聚腺苷酸化信号。 NIH Inua的J.Dean博士在友善的启动子DNA片段中，在交配之前，用PMS/HCG将转化引入小鼠胚胎BDF1雌性小鼠中的小鼠BDF1雌性小鼠。注射的胚胎植入了伪孕的小鼠的生物中。使用从尾巴提取的DNA通过PCR分析检查转化的感染。在四只新生的老鼠中发现了转变。它们的两个（_ <2，> 8）将转化的基因转移到其后代。不仅在卵巢组织中，而且在包括肝，心脏，肺在内的其他组织中检测到相应的转录本。但是仅在卵母细胞和Zona pellucida中观察到蛋白质表达。目前正在研究转基因小鼠的Zona pellucida的异源精子结合能力。