水稻株型突变体rad-1和rad-2的鉴定与功能基因克隆

株型是决定水稻等作物产量的核心因素之一,是品种选育的重要指标。本研究从粳稻品种Asominori的辐射诱变中分离出2个稳定的株型突变体,rad-1和rad-2,它们均表现出苗期弯曲生长、成熟期株高矮化、粒长变短、千粒重降低和产量下降等特征。等位性测验结合连锁分析证实rad-1和rad-2等位,且位于水稻第7染色体上约230kb的范围内。对定位区段的序列分析后确定OsFH5为候选基因,该基因呈现组成型表达,编码水稻Ⅱ型成蛋白。突变体rad-1在OsFH5基因第2个外显子上缺失8个碱基,导致移码;而rad-2则在第14内含子上发生单碱基的变异,发生异常剪切。两类突变最终均导致OsFH5基因翻译提前终止,产生截短的蛋白。相比野生型,rad-1和rad-2在幼苗中OsFH5基因的表达下调。细胞学研究表明,OsFH5基因的功能缺失会导致幼苗叶鞘细胞大小不均,呈现不规则生长,在成熟的颖壳中细胞显著变短。对生长素响应的ARF因子进行表达量检测发现,rad-2中一系列ARF成员表达均显著下调,推测OsFH5极有可能影响了植株对生长素的响应。

Identification and Map-based Cloning of rad-1 and rad-2, Two Rice Architecture Determinant Mutants

Plant architecture is a key element of crop yield and also taken as a criterion in breeding. In this study, two rice architecture determinant mutants, rad-1 and rad-2 were selected from Gamma-radiation induced mutants of rice cultivar Asominori. Each of them shows winding seedlings, dwarf plants and thereby decreased yield. It has been estimated that rad-1 and rad-2 are allelic to each other by allelism test and linkage analysis. After map-based cloning, the responsible gene for the mutations was restricted within a 230 kb region in chromosome 7. Sequence analysis revealed that OsFH5 gene, encoding a forming-like protein altered both in rad-1 and rad-2. An 8-bp deletion in exon 2 in rad-1 and a single nucleotide change in intron 14 in rad-2 made frame-shift reading and produced truncated proteins. OsFH5 gene exhibited constitutive expression in tested tissues. Compared with the wild type, prominent decline of OsFH5 gene was observed in each mutant. In seedlings, loss-function of OsFH5 gene resulted in irregular cell growth in leaf sheath and shorter cells in inner husk, which therefore caused short grain and lower grain weight. By quantitative RT-PCR analysis, several auxin response factors were dramatically down-regulated in rad-2, which suggested that OsFH5 may affect the response to auxin in rice.