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T-DNA标签技术是分离和研究植物功能基因的有效方法,寻找T-DNA插入表型突变体是进一步开展研究的关键所在。文章对以ZH11、ZH15为受体亲本构建的4416份T1代标签系进行了表型鉴定,发现存在拟纯合突变和系内分离突变两种类型,突变表型涉及株高、生育期、叶形、叶色、分蘖力、植株松紧度、穗颈节、穗形、颖花、粒形、类病变、雄性不育、生长极性等14类性状。其中,株高、生育期、叶色、雄性不育有着相对较高的突变频率(超过1%),株高和叶色的突变频率在品种及年度问表现稳定,而生育期、雄性不育波动较大,表明这类性状的表型易受到环境的影响。通过T1、T2连续世代的共分离分析,筛选出3个与穗部或颖花发育相关的T-DNA插入突变体,为分离相关功能基因奠定基础。随机选择42份有表型突变的标签系,通过质粒拯救和TAIL-PCR的方法分离其侧翼序列,从39个标签系中获得40条序列,其中25条为载体序列,14条与水稻基因组有很好的同源性,BlastN分析结果表明T-DNA有优先整合进植物功能基因内部的特性。
T-DNA tagging technology is an effective method to isolate and study plant functional genes. Looking for T-DNA insertion phenotype mutants is the key point for further research. In this study, 4416 T1 generation tag lines constructed with ZH11 and ZH15 as parents were phenotyped and found that there were two types of homozygous mutation and in-line segregation mutation. The mutant phenotype involved in plant height, growth period, leaf 14 traits such as shape, leaf color, tillering ability, plant tightness, panicle neck, spike, spikelet, grain shape, lesion, male sterility and growth polarity. Among them, the plant height, growth period, leaf color and male sterility had a relatively high mutation frequency (over 1%), and the mutation frequency of plant height and leaf color was stable in varieties and annuals, but the growth period, male sterility Fluctuations, indicating that the phenotype of such traits vulnerable to the impact of the environment. Through the co-segregation analysis of T1 and T2 successive generations, three T-DNA insertion mutants related to panicle development or spike development were screened, which laid the foundation for the isolation of related functional genes. Forty-two tagging lines with phenotypic mutations were randomly selected and their flanking sequences were isolated by plasmid rescue and TAIL-PCR. Forty-five sequences were obtained from 39 tag lines, of which 25 were vector sequences and 14 were related to rice genome Very good homology, BlastN analysis results show that T-DNA preferentially integrated into plant functional genes within the characteristics.