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草鱼(Ctenopharyngodon idella)细菌人工染色体(BAC)文库的构建及其特征分析
Alternative TitleConstruction and characterization of two BAC libraries for grass carp (Ctenopharyngodon idella)
Thesis Advisor朱作言
Degree Grantor中国科学院水生生物研究所
Place of Conferral水生生物研究所
Keyword草鱼(Ctenopharyngodon Idella) Bac 文库 高密度尼龙膜
Abstract本研究分别用pCC1BAC/EcoRI 和pCC1BAC/HindIII 载体首次构建了两个草鱼(Ctenopharyngodon idella)基因组细菌人工染色体(BAC)文库。EcoRI 文库含有53000 个克隆。随机挑取150 个BAC 克隆NotI 酶酶切结果表明:插入片段为20kb~265kb,平均插入片段长度为139.7kb,空载率为2 %,插入片段大于100kb的克隆所占的比率为84%。草鱼单倍体基因组的大小约为1000Mb,因此本文库的覆盖率达到草鱼基因组的7.4 倍。从该文库中找到任意单拷贝基因序列的概率为99.94%。HindIII 文库含有52216 个克隆,空载率2.8 %,插入片段平均长度为121.5kb。覆盖率达到草鱼基因组的6.3 倍。以4×2 的阵式将所有文库克隆制成高密度尼龙膜。每张尼龙膜分别对应于4 个384 孔板。该文库可用于草鱼重要经济性状基因克隆、分子标记开发、物理图谱构建、比较基因组研究和全基因组测序奠定基础,并将为草鱼和其它鲤科养殖鱼类的遗传改良提供理论依据和技术支撑。
Other AbstractTwo genomic bacterial artificial chromosome (BAC) libraries were constructed for the grass carp (Ctenopharyngodon idella), using the CopyControl pCC1BAC/EcoRI cloning vector and pCC1BAC/HindIII respectively. The former library consists of about 53000 clones arrayed in 138 384-well microtiter plates. NotI restriction analysis of 150 white clones picked randomly indicated the average insert size of BAC library is 139.7 kb and the frequency of clones without inserts is 2%. Based on the grass carp haploid genome size of 1000 Mb, this library represents approximately 7.4-fold genome coverage. Theoretically, the probability to find any unique sequence in library screening is 99.93%. The later BAC library consists of a total of 52216 clones with an average insert size of 121.5 kb and there are less than 2.8 % of empty clones. This library is estimated to contain approximately 6.3 genome equivalents of the genome of the grass carp. These are the first BAC libraries constructed from this species. High-density filters were prepared, each containing an array of 4 plates in duplicate. Those libraries can be utilized for cloning of genes and molecular markers for selection of strains with economic traits and constructing the physical genome map for whole genome sequencing and comparative genome study, witch will provide profound molecular knowledge and a technology platform to improve the genetic traits of grass carp and other farm cyprinid species.
Document Type学位论文
Recommended Citation
GB/T 7714
刘航. 草鱼(Ctenopharyngodon idella)细菌人工染色体(BAC)文库的构建及其特征分析[D]. 水生生物研究所. 中国科学院水生生物研究所,2008.
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