Studies on the Regulatory Genes of Gonadal Differentiation from the Viewpoint of Utilization in Animal Resource Science.

赠款有效期内取得的代表性成果如下：1)克隆了柴山羊（Capra hircus var Shiba）全长4.9 kb的全基因组SRY （Y染色体性别决定区）DNA并进行了测序。2)转染小鼠Sertoli细胞衍生的TM-4细胞，转染人和小鼠SRY DNA，建立转化细胞系。奇怪的是，p450芳香基因的表达被转染的小鼠SRY DNA诱导。目前正在研究这一细胞系，以确定构成假定的SRY级联的基因。3)从胎儿睾丸cDNA文库中分离得到的牛SRY cDNA具有环状构象。4)克隆山羊c-kit cDNA并测序。结果表明，山羊c-kit的胞内结构域具有一个额外的丙氨酸插入，这在除绵羊外的任何其他物种中都没有被描述过。这种植入物似乎仅限于山羊和绵羊。正在进行更多的尝试来确定插入物的生物学意义。5)在人类性别分化障碍患者中发现了有趣的突变基因。研究是为了阐明位于Y染色体上的基因的功能，并推测其促进了身体的生长。6)对小鼠W^f和W^<sh>突变体表型的详细分析揭示了一些有趣的特征，这些特征可能有助于理解反刍动物c-kit/Sl系统的功能。7)发现了可能与大鼠卵巢上皮细胞凋亡有关的新基因。8)研究了癌基因在卵泡凋亡变化中的作用。研究了细胞周期蛋白在卵母细胞成熟分裂中的作用。10)分析WAP启动子的组织特异性，发现WAP启动子在乳腺以外的许多组织中都有作用。11)通过限制性酶联合处理和PCR分析，选择成功将转基因基因导入基因组的胚胎，试图提高转基因动物的生产效率。少

Representative results obtained during the tenure of the grant are as follows. 1) Coplete genomic SRY (sex determining region on Y chromosome) DNA of Shiba goat (Capra hircus var Shiba) spanning 4.9 kb was cloned and sequenced. 2) Transformed cell lines were established by transfecting TM-4 cells derived from mouse Sertoli cells, with human and mouse SRY DNA.Curiously, as it turned out, the expression of P450aromatese genes was induced by the transfected mouse SRY DNA.This cell line is currently investigated in order to identify the genes which constitute the putative SRY cascade. 3) Bovine SRY cDNA isolated from a fetal testicular cDNA library is confirmed to possess circular conformation. 4) Caprine c-kit cDNA was cloned and sequenced. It turned out that the intracellular domain of caprine c-kit possess one extra alanine insert which have not been described in any other species except sheep so far examined. The insert appears to be limited to goats and sheep. Attempts are being made … More to determine the biological meaning of the insert. 5) Interesting mutant genes were identified in human patients suffering from the disorders of sexual differetiation. Investigations were made to clarify the function of the gene located on Y chromosome and putatively promote the growth of the body. 6) Detailed analysis of phenotypes of W^f and W^<sh> mouse mutants revealed several interesting features which may be useful to approach the understanding of functioning of c-kit/Sl system in ruminants. 7) Novel genes which might be responsible for the apoptosis of lutcal cells in the rat ovary, were identified. 8) Functions of oncognenes in apoptotic changes in the follicles have been investigated. 9) Role of cyclins in maturation division of oocytes has been investigated. 10) Tissue specificity of the WAP promoter was analyzed, and found to be a 'lcaky' promoter functioning in many tissues other than the mammary gland. 11) Attempts were made to improve efficiency of the production of transgenic animals by selecting embryos which successfully incorporated the transgenes into their genome by combined treatment with restrictions cnzymes and PCR analysis. Less