载有抗逆性基因的多枝赖草染色体连锁群的特定TAC克隆的鉴定  

Identification of the Transformation-competent Artificial Chromosome(TAC) Clones Specific to the Linkage Group of Chromosomes Containing Stress-tolerant Genes of Leymus multicaulis

作　　者：张艳[1,2] 赵茂林[2] 孙淼[1,2] 徐粤宇[3] 

机构地区：[1]首都师范大学生命科学学院,北京100037 [2]北京市农林科学院农业生物技术研究中心,北京100097 [3]湖南工程学院,湖南湘潭411104

出　　处：《华北农学报》2008年第5期6-12,共7页Acta Agriculturae Boreali-Sinica

基　　金：国家自然科学基金(30370905,30571135);北京市自然科学基金(5032009)

摘　　要：分别以耐黄矮病耐蚜虫小赖麦附加系Line24、耐盐耐旱小赖麦附加系Line15、耐盐小赖麦易位附加系Line14为材料,以它们各自所附加的含有相应抗逆基因的多枝赖草染色体（简称为抗逆染色体）的6个特定SSR分子标记为探针,通过两轮菌落杂交法筛选多枝赖草基因组可转化人工染色体（TAC）文库,最终得到经过验证的阳性克隆77个,它们分别对应于3个（易位）附加系中所附加抗逆染色体（臂）连锁群的6个特定SSR标记,每一标记所对应的TAC克隆数目变化在5-19个,平均约13个。进一步从来自Line24的抗逆染色体连锁群的同一个探针筛选到的阳性克隆中抽查2个TAC克隆15J18和09I02,通过双色荧光原位杂交技术（Double-color FISH）进行初步定位,直观地证明了文库筛选结果的准确性。为各条抗逆染色体的识别鉴定和相关抗逆基因的精细定位奠定了基础。Leymus multicatdis（2n = 4X = 28, XmXmNsNs）is one of benefit native grass species in Xinjiang, which has many excellent traits such as immune or highly resistant to BYDV（Badey yellow dwarf virus）,and highly tolerant to salt, drought, leanness, pollution, aphids and so on. So it is very important to isolate and clone genes controlling these traits in Leymus multicaulis. In this research, the transformation-competent artificial chromosome （TAC）genomic libraries of Leymus multicaulis were screened through the method of colony blot hybridization in two steps with the probes of specific SSR molecular markers from stress-tolerance chromosomes, which contained stress-tolerant genes of Leymus multicaulis respectively added to wheat alien disomic addition line. Line 24,tolerant to BYDV and aphids;Line 15, tolerant to salt and drought, and wheat alien disomic translocation-addition line; Line 14, tolerant to salt. As a result, a total of 77 positive clones respectively specific to six SSR markers from three stress-tolerance chromosome linkage groups were identified. The accuracy of the screening results was verified visually by physical localization of two TAC clones 15J18 and 09102 from the positive clones specific to a SSR marker of the same linkage group using double-color fluorescent in situ hybridization（Double-color FISH）.The research results will play a pivotal role in identifying individual stress-tolerance chromosome of Leymus multicaulis and constructing high-resolution physical map.

关 键 词：多枝赖草 TAC克隆 二体(易位)附加系 抗逆性 双色荧光原位杂交 

分 类 号：Q781[生物学—分子生物学] Q243