Reporter mice express green fluorescent protein at initiation of meiosis in spermatocytes.
Reporter mice express green fluorescent protein at initiation of meiosis in spermatocytes.
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报告小鼠在精母细胞减数分裂开始时表达绿色荧光蛋白。
DOI：
10.1002/dvg.22830
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Eddy,EdwardM
中科院分区：
文献类型：
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作者：
Brown,PaulaR;Odet,Fanny;Bortner,CarlD;Eddy,EdwardM
利用热休克蛋白2（Hspa 2）基因启动子在精母细胞减数分裂前期I开始时表达绿色荧光蛋白（GFP），构建转基因小鼠。通过原位荧光、免疫组织化学、蛋白质印迹和PCR检测，在四个细胞系中证实了表达。GFP和HSPA 2蛋白的表达和分布共定位于三个系的精母细胞和精子细胞中，但GFP表达在一个系（F46）中是多样化的，存在于减数分裂和减数分裂后生殖细胞的一些克隆中，而不在其他克隆中。荧光激活细胞分选（FACS）用于分离纯化的精母细胞和精子细胞群体。虽然亚硫酸氢盐测序揭示了差异的DNA甲基化模式在启动子区的转基因的杂色表达的GFP线，一个均匀表达的GFP报告线，和theHspa 2基因，这些差异并没有相关的杂色表达。Hspa 2 ‐GFP报告小鼠通过在分离的生精细胞中原位检测GFP，为减数分裂研究提供了一种新的工具。它们将允许对减数分裂和减数分裂后生殖细胞进行分选，以表征分子特征以及GFP表达与阶段特异性生精细胞蛋白和发育事件的相关性。Genesis 52：976-984，2014.© 2014 Wiley Periodicals，Inc.
Transgenic mice were generated using a heat shock protein 2 (Hspa2) gene promoter to express green fluorescent protein (GFP) at the beginning of meiotic prophase I in spermatocytes. Expression was confirmed in four lines byin situfluorescence, immunohistochemistry, western blotting, and PCR assays. The expression and distribution of the GFP and HSPA2 proteins co‐localized in spermatocytes and spermatids in three lines, but GFP expression was variegated in one line (F46), being present in some clones of meiotic and post‐meiotic germ cells and not in others. Fluorescence activated cell sorting (FACS) was used to isolate purified populations of spermatocytes and spermatids. Although bisulfite sequencing revealed differences in the DNA methylation patterns in the promoter regions of the transgene of the variegated expressing GFP line, a uniformly expressing GFP reporter line, and theHspa2gene, these differences did not correlate with variegated expression. TheHspa2‐GFP reporter mice provide a novel tool for studies of meiosis by allowing detection of GFPin situand in isolated spermatogenic cells. They will allow sorting of meiotic and post‐meiotic germ cells for characterization of molecular features and correlation of expression of GFP with stage‐specific spermatogenic cell proteins and developmental events. genesis 52:976–984, 2014. © 2014 Wiley Periodicals, Inc.