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DNA sequencing system from histopathological sections

组织病理切片 DNA 测序系统

基本信息

批准号：
10556068
负责人：
KON Yasuhiro
金额：
$ 8万
依托单位：
HOKKAIDO UNIVERSITY
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

项目摘要

In the present study, DNA sequencing system was established from histopathological sections by using microdissection procedure, and several mutant mice were investigated by morphological and molecular biological approaches. Mutant mice investigated were the following ; 1) urethan-induced lung tumor, 2) renin knockout mice, 3) transgenic mice overexpressing GM2 activator protein, 4) hereditary cataract mice, 5) female genital tumor mice creating by transgenic strategy and 6) male sterile mice. Additionally, the novel genomic polymorphisms were detected by the host genome subtraction method using Marek's Disease virus infected cell lines.During the investigation of spermatogenesis, MRL mouse was established as a new strain carrying the genes of meiotic metaphase-specific apoptosis and heat-shock resistant spermatocytes. Surprisingly, this strain showed temporarily oocyte-like cells in testis during postnatal periods. These results assumed that spermatocyte possessing heat-shock resistance in MRL mice differentiates into oocytes-like cells before the completion of meiotic checkpoint in Sertoli cells, and then progresses to apoptotic figure of meiotic metaphase after it's completion. In order to identify the genes associating meiotic metaphase-specific apoptosis and heat-shock resistance spermatocytes, the differential display was performed between the testis cDNAs of C57BL/6 and MRL mice, detecting several novel clones. Especially, it was suggested by chromosomal linkage analysis of 200 backcross progenies that the gene carrying metaphase-specific apoptosis was mapped at 100.0 cM on chromosomal 1.

本研究采用显微切割技术建立了组织病理切片的DNA测序系统，并通过形态学和分子生物学方法对几种突变小鼠进行了研究。所研究的突变小鼠如下：1）β-内酰胺酶诱导的肺肿瘤，2）肾素敲除小鼠，3）过表达GM 2激活蛋白的转基因小鼠，4）遗传性白内障小鼠，5）通过转基因策略产生的雌性生殖器肿瘤小鼠和6）雄性不育小鼠。在精子发生的研究中，建立了一个新的MRL小鼠品系，该品系携带有减数分裂中期特异性凋亡基因和热休克抗性基因。令人惊讶的是，该菌株在出生后的睾丸中暂时显示出卵母细胞样细胞。这些结果表明，MRL小鼠具有热休克抗性的精母细胞在支持细胞减数分裂检查点完成前，先分化为类卵母细胞，在检查点完成后，再向减数分裂中期的凋亡样细胞发展。为了鉴定精母细胞减数分裂中期特异性凋亡与热休克抗性相关的基因，对C57 BL/6和MRL小鼠睾丸cDNA进行差异显示，发现了几个新的克隆。特别是对200个回交后代的染色体连锁分析表明，携带中期特异性凋亡的基因定位在第1染色体上100.0cM处。