Crop Breeding and Cultivation Research Institute, Shanghai Academy of Agricultural Sciences, Shanghai, China.
PLoS One. 2012;7(8):e43026. doi: 10.1371/journal.pone.0043026. Epub 2012 Aug 24.
Foods high in resistant starch (RS) are beneficial to prevent various diseases including diabetes, colon cancers, diarrhea and chronic renal or hepatic diseases. Elevated RS in rice is important for public health since rice is a staple food for half of the world population. A japonica mutant 'Jiangtangdao 1' (RS = 11.67%) was crossed with an indica cultivar 'Miyang 23' (RS = 0.41%). The mutant sbe3-rs that explained 60.4% of RS variation was mapped between RM6611 and RM13366 on chromosome 2 (LOD = 36) using 178 F(2) plants genotyped with 106 genome-wide polymorphic SSR markers. Using 656 plants from four F(3:4) families, sbe3-rs was fine mapped to a 573.3 Kb region between InDel 2 and InDel 6 using one STS, five SSRs and seven InDel markers. SBE3 which codes for starch branching enzyme was identified as a candidate gene within the putative region. Nine pairs of primers covering 22 exons were designed to sequence genomic DNA of the wild type for SBE3 and the mutant for sbe3-rs comparatively. Sequence analysis identified a missense mutation site where Leu-599 of the wild was changed to Pro-599 of the mutant in the SBE3 coding region. Because the point mutation resulted in the loss of a restriction enzyme site, sbe3-rs was not digested by a CAPS marker for SpeI site while SBE3 was. Co-segregation of the digestion pattern with RS content among 178 F(2) plants further supported sbe3-rs responsible for RS in rice. As a result, the CAPS marker could be used in marker-assisted breeding to develop rice cultivars with elevated RS which is otherwise difficult to accurately assess in crops. Transgenic technology should be employed for a definitive conclusion of the sbe3-rs.
富含抗性淀粉 (RS) 的食物有益于预防各种疾病，包括糖尿病、结肠癌、腹泻和慢性肾或肝疾病。由于大米是全球一半人口的主食，因此大米中 RS 的升高对公众健康很重要。粳稻突变体“江道 1 号”（RS=11.67%）与籼稻品种“绵阳 23 号”（RS=0.41%）杂交。在 178 株 F2 代植物中，用 106 个全基因组多态性 SSR 标记进行基因型分析，将解释 RS 变异 60.4%的突变体 sbe3-rs 定位在第 2 染色体上的 RM6611 和 RM13366 之间（LOD=36）。利用来自四个 F3:4 家系的 656 株植物，通过一个 STS、五个 SSR 和七个 InDel 标记，将 sbe3-rs 精细定位到 InDel 2 和 InDel 6 之间的 573.3 Kb 区域。淀粉分支酶编码 SBE3 基因被鉴定为该假定区域的候选基因。设计了 9 对引物，涵盖 22 个外显子，用于比较野生型 SBE3 和突变型 sbe3-rs 的基因组 DNA 测序。序列分析确定了一个错义突变位点，野生型的亮氨酸 599 突变为突变型的脯氨酸 599。由于点突变导致一个限制酶位点的丢失，sbe3-rs 不能被 SpeI 位点的 CAPS 标记消化，而 SBE3 可以。在 178 株 F2 植物中，消化模式与 RS 含量的共分离进一步支持 sbe3-rs 是水稻 RS 的原因。因此，该 CAPS 标记可用于标记辅助选择，以开发 RS 升高的水稻品种，否则在作物中很难准确评估。应采用转基因技术对 sbe3-rs 进行明确的结论。
