| 题名 | 中国穿山甲细菌人工染色体文库的构建及其特征鉴定 |
| 作者 | 车骏 |
| 学位类别 | 硕士 |
| 答辩日期 | 2007-07 |
| 授予单位 | 中国科学院研究生院 |
| 授予地点 | 北京 |
| 导师 | 杨凤堂 |
| 关键词 | 中国穿山甲 细菌人工染色体（BAC）文库 荧光原位杂交 细胞遗传学 |
| 其他题名 | Construction and characterization of a Bacterial Artificial Chromosome (BAC) library of the Chinese pangolin (Manis pentadactyla) |
| 学位专业 | 细胞生物学 |
| 中文摘要 | 鳞甲目 （Pholidota）作为真兽亚纲18个现生目之一，是理解哺乳动物系统分类和进化的一个关键类群。穿山甲的形态十分特殊，它的系统发生地位历来就是人们关注的对象，搞清穿山甲的系统发生地位将有助于理解和重建整个哺乳动物的系统发生过程。目前，中国穿山甲已被列入“哺乳动物基因组计划”24个待测序物种其中之一。细胞遗传学的研究表明中国穿山甲的核型具有多态性。 细菌人工染色体 （BAC）文库目前已成为基因组学研究领域的最为重要的研究工具之一。各种动植物细菌人工染色体文库的逐渐丰富为后基因组时代比较基因组研究奠定了良好的基础。中国穿山甲基因组DNA文库的构建将有力地促进穿山甲染色体演化、哺乳动物比较基因组学及其系统发育学的研究。 本研究在世界上首次成功地构建了一个具有约8倍基因组覆盖率的中国穿山甲细菌人工染色体文库。该基因组文库含有208,512个克隆，储存在543块384孔板中。对随机选取的171个克隆的Not1酶切和脉冲场电泳结果分析显示: 64.5%的克隆插入片段大小介于100-149 Kb之间， 75.6%的克隆插入片段大于100kb。平均插入片段大小为111.4Kb; 非重组克隆 （无插入片段）的比率为9.9%。同时，我们通过荧光原位杂交把105个BAC克隆定位到了中国穿山甲染色体上的特异区域, 构建了中国穿山甲的BAC细胞遗传学图谱。初步的染色体定位结果表明，BAC克隆在基因组中的分布没有偏向性。 中国穿山甲BAC文库的成功构建，丰富了哺乳动物基因组文库资源。该文库除有利于比较基因组研究之外，也为深化保护遗传学研究奠定了基础。BAC细胞遗传学图谱的构建，将为中国穿山甲全基因组测序中产生的序列重叠群在染色体上的定向和定位奠定基础。 |
| 英文摘要 | Pangolins, representing one of the 18 extant Eutheria orders, occupy an important evolutionary status and deserve extensive research. With some extremely specified morphological characters, the phylogenetic position of pangolins has attracted the interests from many researchers. The elucidation of the position of pangolins on mammalian phylogenetic tree is important for the reconstruction and understanding of the evolutionary history of mammals. Recently, Chinese pangolin (Manis pentadactyla) has been enlisted in the “mammalian genome project”. Previous cytogenetic studies show that there is karyotypic polymorphism in Chinese pangolins. Nowadays Bacterial Artificial Chromosome (BAC) library has become one of the most important tools in genomic study. More and more BAC libraries of different species of animals and plants have laid the foundation for comparative genomics studies in the post-genomics era. The construction of a BAC library for the Chinese pangolin is essential for mammalian comparative genomics and a better understanding of the mechanism underlying the karyotypic evolution of pangolins. Here I have constructed an 8 times genomic-coverage BAC library for the Chinese pangolin. The library consists of 208,512 clones that are stored in 534 × 384-well plates. Pulse field gel eletrophoresis analysis of randomly selected 171 BAC clones digested with Not1 restriction enzyme shows that 64.5% of clones have the insert size between 100-149 kb, while 75.6% of the clones have the insert size larger than 100 kb. The average insert size is 111.4 kb, and the ratio of non-recombinant clones is 9.9%. In addition, 105 BAC clones were mapped onto specifc chromosomal regions of Chinese pangolin chromosomes by fluorescence in situ hybridization (FISH). This BAC library can serve as a valuable resource for conservational genetic study as well as comparative genomic study. The cytogenetic map of BAC clones will be useful for the chromosomal orientation and positioning of sequence contigs generated by the proposed whole-genome sequencing project of Chinese pangolin as well as for the integration of cytogenetic map with other genetic or physical maps. |
| 语种 | 中文 |
| 公开日期 | 2010-10-14 |
| 内容类型 | 学位论文 |
| 源URL | [http://159.226.149.42:8088/handle/152453/6120]  |
| 专题 | 昆明动物研究所_其他 |
| 推荐引用方式 GB/T 7714 | 车骏. 中国穿山甲细菌人工染色体文库的构建及其特征鉴定[D]. 北京. 中国科学院研究生院. 2007. |
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