FUNCTION OF CHROMOSOMAL REGIONS FOR GENOME STABILITY

利用最近发展的TAR(转换相关重组)克隆技术，可以利用完整或部分序列信息直接从复杂的基因组中分离出特定的染色体区域和基因作为大的线性或环状YAC。在去年，一项工作的重点是进一步优化焦油克隆，使这项技术在科学界得到更广泛的应用。此外，成功地分离了四个人类基因的完整拷贝，包括转移抑制基因KAI1、父系表达基因Ubiq和端粒酶逆转录酶hTERT基因，用于进一步的功能研究。TAR克隆技术也被应用于人类染色体着丝粒区域的分离和鉴定。作为这项研究的一部分，我们测定了人类Y染色体着丝粒区域的完整序列。该着丝粒片段含有约140kb的αDNA，用于构建人小圆形人工染色体缺失端粒(HAC)。当它被导入人体细胞时，它稳定地保持在每个细胞1-2个拷贝。HAC克隆系统的发展为人类基因治疗提供了新的机遇，并可能对我们理解人类染色体分离的机制产生深远的影响。

Using the recently developed TAR (Transformation-Associated Recombination) cloning technique, it is possible to directly isolate specific chromosomal regions and genes from complex genomes as large linear or circular YACs using a complete or a partial sequence information. During last year a work was focused on further optimizing of TAR cloning to make the technology more widely used in the scientific community. In addition, entire copies of four human genes including metastasis-suppressor gene KAI1, a paternally expressed gene Ubiq and the telomerase reverse transcriptase hTERT gene were successfully isolated for further functional studies. TAR cloning technique has been also applied for isolation and characterization of centromeric regions of human chromosome. As a part of this study a complete sequence of centromeric region of the human Y chromosome was determined. This centromeric fragment containing ~140 kb of alphoid DNA was used for construction of small circular Human Artificial Chromosome lacking telomeres (HAC). When transfected into human cells, it is stably maintained at 1-2 copies per cell. Development of the HAC cloning system provides new opportunities for human gene therapy and could have profound effects on our understanding of mechanisms of human chromosome segregation.