24种香稻品种甜菜碱醛脱氢酶2基因突变位点的分析及分子标记开发

Development of Molecular Markers Used to Identify Two Types of Fragrant Rice and Analysis of Mutation Sites of BADH2Gene in 24 Varieties of Fragrant Rice

根据香型与非香型水稻甜菜碱醛脱氢酶2基因(badh2)在第2、第4内含子、第7外显子3处序列差异和第2外显子1处序列差异,分别设计了两类检测badh2第7和第2外显子突变的功能性分子标记引物M7和M2;利用两类引物,分别对属于第7外显子突变的香型水稻W香99075和第2外显子突变的香型水稻武香14、非香型水稻以及两种香稻分别与非香稻杂交的F1植株基因组DNA进行PCR检测后发现,M7和M2引物完全能够分别被用于以第7和第2外显子突变的香稻作为亲本,进行分子标记辅助培育香稻新品种的研究。M7引物综合考虑了badh2内含子和外显子两方面突变情况而设计的。以非香稻261S、分别发生第7和第2外显子突变的香稻品种W香99075和武香14为对照,使用M7和M2引物,对本实验室收集的另外22份香稻品种进行badh2突变位点检测,结果可将这些香稻分为badh2第2外显子突变类型、第7外显子突变类型和外显子未发生突变类型,同时明确了大多目前在上海等周边地区种植的香稻品种的badh2所属的突变位点。开展本研究为利用分子标记辅助选育香型水稻新品种研究奠定了重要的基础。

Functional molecular markers M7 and M2 have been developed based on the DNA sequence differences of badh2 between fragrant rice varieties and non-fragrant varieties in intron2, intron 4, exon7 and exon 2 respectively. PCR analyses on genome DNA of exon7-mutated fragrant rice Wxiang 99075, exon2-mutated fragrant rice Wux- iangl4, non-fragrant rice 261S and the F1 plants by M7 and M2 showed that M7 and M2 could be absolutely used to the molecular marker-assisted rice breeding experiments when exon7-mutated and exon2-mutated fragrant rice varie- ties are used as parents. The design of M7 primers took mutations both in exons and intrones into account. Moreover, taking 261S, Wxiang 99075 and Wuxiangl4 as controls, the mutation sites of badh2 in 22 fragrant rice varieties were analyzed, it was showed that fragrant rice varieties could be classified into 3 types ： exon 2-mutated fragrant rice, exon 7-mutated fragrant rice and non-exon-mutated fragrant rice. At the same time, the mutation sites of badh2 in the main fragrant rice varieties which are grown in Shanghai and the surrounding areas have been verified. This research laid an important foundation for molecular marker-assisted selection for novel fragrant rice.