HUMAN DISTAL EQ DNA--ANALYSIS WITH YACS

人类细胞遗传学部分DNA的系统分析 Xq 24-Xq 28是利用酵母人工 染色体（YACs）。 一些基因被包括在病变引起的 单基因疾病 这些基因可以完整地分离出来， 为医疗应用提供长期潜力。 研究 也可以是一个试点项目， 染色体区域 启动工作的测试库的总 已经开发了YAC中的人DNA，用于筛选 兴趣 第二个图书馆正在组织从仓鼠人类 体细胞杂种，其中Xq 24-Xq 28区域是唯一已知的 人类DNA;该文库现在含有10 Mb人类特异性DNA 在60个克隆中，并将在 项目任期将开始。 在混合体的文库中， 含有人类DNA的DNA的基因的出现频率 的细胞DNA（1/250），并且已经产生了两个YAC 其与单个Xq 28探针杂交;这两个克隆重叠 在他们的限制性模式，这表明YAC含有 未重排的DNA片段 一些可用的探针将被 用于筛选两个库，并辅助映射技术 将用于搜索重叠。 重叠群会特别 寻找MN 12-st 14和葡萄糖-6-磷酸脱氢酶 （G6 PD）-Xq 28的工厂VIII色盲区域。 选择 将分析区域的转录位置和 某些重复序列，通过限制性位点 cDNA文库的定位和筛选。 最后，一个完整的形式 将寻求G6 PD基因，以评估需求 用于转染细胞中的表达和mRNA加工。

Systematic analysis of DNA from portions of the human cytogenetic bands Xq24-Xq28 is proposed, based on the use of yeast artificial chromosomes (YACs). Some genes are included in which lesions cause monogenic disease. Those genes could be isolable in intact form, providing long-term potential for medical applications. The study could also be a pilot project for the complete analysis of a chromosomal region. To initiate the work a test library of total human DNA in YACs has been developed to be screened for clones of interest. A second library is being organized from a hamster-human somatic cell hybrid in which the Xq24-Xq28 region is the only known human DNA; that library now contains 10 Mb of human-specific DNA in 60 clones, and will be expanded to l-hit (30 Mb) by the time the project tenure would begin. In the library from the hybrid, clones containing human DNA occur with a frequency equal to their fraction of the cellular DNA (1/250), and have already yielded two YACs which hybridize to a single Xq28 probe; those two clones overlap in their restriction patterns, suggesting that the YACs contain unrearranged DNA segments. A number of available probes will be used to screen the two libraries, and subsidiary mapping techniques will be used to search for overlaps. Contigs will be particularly sought for the MN12-st14 and glucose-6-phosphate dehydrogenase (G6PD)-Factory VIII-color blindness regions of Xq28. Selected regions will be analyzed for the locations of transcribed and of certain repetitive sequences, by a combination of restriction site mapping and screening of cDNA libraries. Finally, an intact form of the G6PD gene will be sought in order to assess the requirements for expression and mRNA processing in transfected cells.