水稻T-DNA插入突变体库的筛选及遗传分析

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T，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间表现稳定，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。

Screening for and Genetic Analysis on T-DNA-inserted Mutant Pool in Rice

T-DNA tagging technique has provided a powerful strategy for identifying new functional genes in plants, and the key for success is the discovery of T-DNA-inserted mutants with changed phenotype. In this study, we screened 4 416 rice T1 tagged lines generated by enhancer trap system integrated with GLL4/VP16-UAS elements from two transformed parents, ZH11 and ZH15. We found many lines showed obvious morphological mutations, including two typesake-homozygous mutation and separating mutation. The mutation phenotype was related to 14 kinds of trait such as plant height, heading date, leaf shape, leaf color, tiller number, panicle shape, spikelet number, grain shape, disease-like mutation, male sterility, awn, and so on. Among them, plant height, heading date, leaf color and male sterility had a comparatively high mutation frequency (over 1%). The mutation frequency of plant height and leaf color had no significant change between different years or transformed parents, but the frequency of heading date and male sterility varied greatly, suggesting that environment had a great effect on the expression of latter two traits. By conducting continuously co-segregating analyses in T1 and T2 generation, we identified 3 T-DNA-inserted mutants with malformed panicle or spikelets, which would provide a base for cloning correlative functional genes. At the same time, we selected randomly 42 lines with mutation phenotype and obtained 40 flanking sequences from 39 tagged lines by plasmid rescue or TAIL-PCR, of which, 26 were vector backbone sequence, 14 had good identity to rice genome sequence. The BlastN result showed the T-DNA preferentially integrated into protein-coding region in plants.