甘蓝型油菜基因组文库的构建及与硼高效基因相连锁克隆的筛选(英文)

Construction of a Brassica napus Bacterial Artificial Chromosome Library and Identification of Clones Linked to Boron Efficiency Gene

以甘蓝型油菜宁RS 1为材料 ,构建了含有 82 94 4个克隆的甘蓝型油菜的BAC基因组文库。从文库中随机挑取克隆进行DNA长度检测 ,BAC克隆平均插入片段大小为 80kb ,覆盖甘蓝型油菜基因组的 5 1倍。随机挑取 10 8克隆进行继代培养 10 0代 ,分离质粒酶切检测表明不存在插入片段丢失现象 ,表明该文库的克隆在大肠杆菌中稳定存在 ;以与硼高效基因相连锁的分子标记拟南芥cDNA克隆pa2 8为探针对文库进行了筛选 ,共获得 13个阳性克隆 ,进一步研究表明 ,3个克隆 13E3、32G8、186G2 0与BE1基因相连锁。

A bacterial artificial chromosome (BAC) library was constructed for a Brassica napus cultivar: Ning RS 1. The library contains 82944 clones with 80kb DNA insert size in average representing equivalents of 5 1 Brassica napus haploid genome. The BAC DNA can maintain stably in E. coli after about 100 generations of culture. pa28, an Arabidopsis EST clone which linked to a boron efficiency gene BE1 in B.napus , was used as probe to screen the library: consequently, 13 positive clones were identified. These positive clones were divided into four groups. Three of 13 clones were identified to link to the BE1 locus by Southern analysis.