Analysis of chromatin structure and gene function of chicken sex chromosomes

鸡的性染色体组成为母鸡ZW，公鸡ZZ。大约2/3的W染色体和Z染色体的末端区域是异染色质。两个W染色体特异的重复DNA序列，XhoI家族和EcoRI家族，是W异染色质的主要成分，但Z末端异染色质的性质尚不清楚。本研究1)从母鸡基因组文库中获得了一个全长约13kb的基因组克隆pCHZTH8。荧光原位杂交(FISH)、Southern杂交和核苷酸序列分析表明，该克隆序列是位于Z-末端异染色质的大卫星DNA的一部分，该序列的大部分是鸡属所特有的。2)虽然大约有20种表型被定位在Z染色体上，但分子水平上的信息尚不清楚。通过基因克隆和FISH技术，我们证明了ZOV3(属于免疫球蛋白超家族的基因)和IREBP(鸡的铁反应元件结合蛋白同源基因)位于Z染色体的不同臂上。3)除上述两个重复的家系外，W染色体特异的DNA序列未知。通过激光显微切割和随机PCR扩增，构建了W染色体特异的基因组文库。获得了两个含有W-特异性非重复序列的克隆(CW01和CW50)。用CW01和CW50筛选母鸡基因组文库，得到两个约25kb的基因组克隆，分别含有CW01和CW50序列。

Constitution of chicken sex chromosomes is ZW for female and ZZ for male. About 2/3 of the W chromosome and a terminal region of the Z chromosome are heterochromatic. Two W-chromosome-specific repetitive DNA sequences, XhoI family and EcoRI family, are major constituents of the W heterochromatin, but nature of the Z-terminal heterochromatin is unknown. In this study, 1) a genomic clone pCHZTH8 containing about 13kb genomic fragment was obtained from the female chicken genomic library. Fluorescence in situ hybridization (FISH), Southern blot hybridization and nucleotide sequencing revealed that this cloned sequence was a part of the macrosatellite DNA localized in the Z-terminal heterochromatin and that most part of this sequence was specific to the genus Gallus. 2) Although about 20 phenotypes are mapped to the Z chromosome, molecular-level information has not been available. We demonstrated by cDNA cloning and FISH that ZOV3 (a gene belonging to the immunoglobulin superfamily) and IREBP (chicken homolog of iron responsive element binding protein) were localized on different arms of the Z chromosome. 3) Except for the above two repetitive families, W chromosome-specific DNA sequences are unknown. We constructed a W chromosome-specific genomic library by applying laser microdissection and random PCR amplification. Two clones containing W-specific non-repetitive sequences (CW01 and CW50) were obtained. A female chicken genomic library was screened with CW01 and CW50, and genomic clones convering two genomic regions of about 25-kb each, containing the CW01 and CW50 sequences, respectively, were obtained.