Construction of a Bacterial Artificial Chromosome Library of TM-1, a Standard Line for Genetics and Genomics in Upland Cotton
Construction of a Bacterial Artificial Chromosome Library of TM-1, a Standard Line for Genetics and Genomics in Upland Cotton
复制标题
DOI：
10.1111/j.1744-7909.2008.00773.x
复制
发表时间：
2009-01-01
影响因子：
11.4
通讯作者：
Zhang, Tian-Zhen
中科院分区：
文献类型：
--
作者：
Hu, Yan;Guo, Wang-Zhen;Zhang, Tian-Zhen
构建了高地棉遗传和基因组标准系陆地棉TM-1的细菌人工染色体（BAC）文库。该文库包含147456个克隆，平均插入片段大小为122.8kb，范围为97 ~ 240 kb。约96.0%的克隆具有大于100 kb的插入片段。因此，基于AD基因组大小为2425 Mb，该文库理论上代表7.4个单倍体基因组当量。将克隆储存在384 384（-）孔板中，并排列成多重池用于快速和可靠的文库筛选。利用23个SSR标记、3个序列相关扩增多态性标记和1对纤维发育相关基因的引物，通过4轮聚合酶链反应对文库进行BAC筛选。相应地，总共鉴定出92个阳性BAC克隆，每个SSR标记平均4个阳性克隆，范围从1到8个命中。此外，根据遗传图谱，这些SSR标记已定位于第12号染色体（A12）和第26号染色体（D12），因此这些BAC克隆有望用作这两条同源染色体的物理作图的种子，从而基于图位克隆数量性状基因座或与重要农艺性状相关的基因。
A bacterial artificial chromosome (BAC) library was constructed for Gossypium hirsutum acc. TM-1, a genetic and genomic standard line for Upland cotton. The library consists of 147 456 clones with an average insert size of 122.8 kb ranging from 97 to 240 kb. About 96.0% of the clones have inserts over 100 kb. Therefore, this library represents theoretically 7.4 haploid genome equivalents based on an AD genome size of 2 425 Mb. Clones were stored in 384 384(-) well plates and arrayed into multiplex pools for rapid and reliable library screening. BAC screening was carried out by four-round polymerase chain reactions using 23 simple sequence repeats (SSR) markers, three sequence-related amplified polymorphism markers and one pair of primers for a gene associated with fiber development to test the quality of the library. Correspondingly, in total 92 positive BAC clones were identified with an average four positive clones per SSR marker, ranging from one to eight hits. Additionally, since these SSR markers have been localized to chromosome 12 (A12) and 26 (D12) according to the genetic map, these BAC clones are expected to serve as seeds for the physical mapping of these two homologous chromosomes, sequentially map-based cloning of quantitative trait loci or genes associated with important agronomic traits.