马铃薯栽培品种‘合作88’细菌人工染色体文库的构建与评价  

Construction and Characterization of a Bacterial Artificial Chromosome Library of Potato Cultivar C88

作　　者：杨煜[1] 杨晓慧[1] 李灿辉[2] 郭晓[1] 单伟伟[1] 马伟清[1] 黄三文[3] 李广存[1,3] 

机构地区：[1]山东省农业科学院蔬菜花卉研究所,山东省设施蔬菜分子生物学重点实验室,济南250100 [2]云南师范大学生命科学学院,昆明650500 [3]中国农业科学院蔬菜花卉研究所,北京100081

出　　处：《园艺学报》2015年第2期361-366,共6页Acta Horticulturae Sinica

基　　金：国家‘863’计划项目(2013AA102603-4);国家科技支撑计划项目(2012BAD02B05-10);山东省良种工程农业生物资源创新利用项目(PTBR2013);公益性行业(农业)科研专项(201303018-03);山东省农业科学院青年科研基金项目(2014QNZ03)

摘　　要：以四倍体马铃薯栽培品种‘合作88’培养12~15 d苗龄的幼嫩叶片为试材,提取高分子量基因组DNA,利用限制性内切酶HindⅢ部分酶切后回收100~300 kb片段,连接到载体Copy ControlTM p CC1BACTM的HindⅢ位点上,构建了细菌人工染色体(BAC)文库。该文库约由850 000个克隆组成,空载率约2.08%,保存在1 700个混合池中,克隆插入片段平均大小约为90 kb,覆盖单倍型马铃薯基因组约85倍。随机挑取6个BAC克隆连续培养5 d,提取DNA进行HindⅢ完全酶切检测,确认不同培养时间的BAC克隆的指纹图谱完全一致,表明文库较好的稳定性。BAC文库的构建为马铃薯重要农艺性状基因克隆、基因组测序以及比较基因组研究等奠定了基础。A bacterial artificial chromosome library of potato(Solanum tuberosum L.)was constructed for‘C88',a tetraploid potato cultivar conferring resistance to late blight,PVX and PVY,using BAC vector Copy ControlTM p CC1. Fresh leaf tissues from 12–15 day-old seedlings were used to isolate high molecular weight genomic DNA,and 100–300 kb partially HindⅢ digested DNA fragments were selected and ligated into the HindⅢ site of p CC1. The library consists of 850 000 clones and stored in 1 700 pools. The average insert size of BAC clones is about 90 kb and the empty clones consist of about 2.08%. The library is equivalent to 85 haploid potato genomes. Six randomly picked clones showed stable HindⅢ fingerprints after 5 days' successive culture,indicating that the clones in the library are stable. The library will facilitate gene cloning,sequencing and comparative genomics research of potato.

关 键 词：马铃薯 细菌人工染色体文库 抗病性 

分 类 号：S532[农业科学—作物学]