早花基因(FT)介导的烟草快速回交改良研究  

Tobacco Rapid Backcross Improvement Mediated by Arabidopsis Flowering Gene FT

作　　者：常爱霞[1] 郭利杰[1,2] 刘旦[1] 罗成刚[1] 王林嵩[2] 冯全福[1] 王兰[2] 

机构地区：[1]中国农业科学院烟草研究所,山东青岛266101 [2]河南师范大学生命科学学院,河南新乡453000

出　　处：《中国农业科学》2015年第13期2508-2517,共10页Scientia Agricultura Sinica

基　　金：中国烟草总公司科技重点项目(110201302003)

摘　　要：【目的】针对常规回交育种周期较长的问题,以烟草为模式植物,探索快速回交改良作物的有效方法。【方法】以对TMV免疫的烤烟品种Fc8作为抗性供体亲本,以综合性状优良但感TMV的中烟100为受体亲本进行回交改良及种质创新。首先从拟南芥中克隆早花FT,将其连接到植物表达载体P22上,并转入农杆菌中,采用叶盘法转化烟草Fc8,创建含FT的Fc8阳性植株。阳性植株分别与中烟100杂交,根据F1后代群体中早花基因的分离比例,鉴定筛选含单拷贝FT的阳性植株。接下来,从含单拷贝FT的阳性植株的F1群体中选择早花植株,与中烟100回交,根据回交分离群体中FT早花基因与TMV抗性N基因的分离比例,筛选FT与N基因不连锁的阳性植株。选择该阳性植株与中烟100连续回交。在每一回交分离世代中,选择早花且含有TMV抗性标记的植株连续回交,当后代遗传背景大部分与中烟100相近,并保留TMV抗性时选择非早花且含N基因标记的植株,鉴定无FT、Ubi启动子、Nos终止子和Hyg筛选标记等转基因元件后,连续自交纯化,筛选改良的创新种质。【结果】通过转基因技术共获得8株含FT的Fc8阳性植株,经鉴定获得一个含单拷贝FT,且FT和TMV抗性N基因不连锁的Fc8植株。用该植株作为TMV抗性的供体亲本回交改良中烟100,一年多时间已回交加代到BC4F1,比常规回交育种时间缩短450—500 d。早花基因在杂交、回交各世代均能稳定表达,早花植株在苗龄50—60 d现蕾开花,早花植株现蕾开花时,一般有3—4片叶,株高平均约35 cm,同期非早花植株高约27 cm,仍然处于营养生长阶段。非早花植株在苗龄130—140 d现蕾开花,平均株高约120 cm,叶片数18—20片。FT能使植株花期提前70—90 d,缩短世代周期约一半时间;早花加代结合TMV分子标记筛选得到了BC1、BC2、BC3代TMV抗性烟草新种质。【结论】借助FT介导的早花及抗TMV基因分子标记,能有【Objective】In order to shorten the conventional back-cross breeding cycle, a rapid back-cross way was studied to improve crops using tobacco as a model plant.【Method】Using flue-cured tobacco variety FC8, which is immune to TMV, as the resistant donor parent, and flue-cured tobacco variety Zhongyan 100, which is susceptible to TMV but with good comprehensive properties, as the receptor parent, the following experiments were conducted. First, the Arabidopsis thaliana FT gene was cloned and connected to plant expression vector P22, which was then transferred into Agrobacterium, and leaf-disc method was adopted for gene transformation to create FC8 positive plants containing FT. These positive plants were hybridized with Zhongyan 100 respectively, and according to the separation ratio between early-flowering and non early-flowering plants in F1 progeny, the positive plants containing single FT gene were identified. Second, the early-flowering plants were selected from the target F1 progeny to backcross with Zhongyan 100. According to the separation ratio of FT and N gene（TMV resistance gene） marker in BC1 progeny, the positive plants containing single FT gene and in which FT gene was not linked to N gene were identified. Next, continuous backcross with Zhongyan 100 was conducted using the specific positive plants, and plants with FT and N gene marker at each earlier backcross generation were selected. In the final generation, the plants containing N gene marker but no FT marker were selected, and identified whether they contained genetically modified components such as FT, Ubi promoter, Nos terminator and Hyg selection marker. Plants free of those components were continuously self-crossed to make N gene isozygoty. 【Result】A total of 8 positive plants containing FT were obtained by transgenic technology, one of which was identified to contain single FT gene, and which FT gene was not linked to TMV resistance gene. Using it as the early-flowering TMV resistance donor parent and Zhongyan100 as the recurr

关 键 词：烟草 FT 快速回交 改良 

分 类 号：S572[农业科学—烟草工业]