无抗性选择标记转基因软米和糯稻新品系的选育及中间试验

Breeding and Field Performance of Novel Soft and Waxy Transgenic Rice Lines without Selectable Markers

直链淀粉含量是影响稻米品质的最主要因素之一。为培育直链淀粉含量较低的软米和糯稻新品系,在经共转化法获得的转反义Wx基因水稻自交后代中筛选获得了不含抗性选择标记基因的纯合体。对这些纯合转基因水稻的分析表明,其未成熟种子胚乳中Wx基因表达水平和成熟种子中Wx蛋白量较未转化对照有不同程度的降低。转基因水稻成熟种子中直链淀粉含量较未转化对照有不同程度的下降,2个转基因系降低至10%左右,相当于软米的直链淀粉含量,1个转基因系降到了2%以下,形成了糯稻新品系。转基因稻米中直链淀粉含量降低后,其胶稠度相应变软,而糊化温度未发生明显改变。田间试验结果表明,转基因水稻的大部分农艺性状与受体品种武香粳9号无显著差异。因此,本研究成功培育了无抗性选择标记且淀粉品质改良的转反义Wx基因水稻新品系。

Amylose content is one of the important factors affecting rice cooking and eating quality, and controlled by the Waxy （Wx） gene which encoding a granule-bound starch synthase. High amylose levels are usually associated with poor cooking and eating quality. In our prevrious studies, in order to generate transgenic rice varieties with improved quality and without of the selectable marker, the antisense Wx gene was co-transformed together with a selectable marker gene, the hygromycin-resistant gene （HPT）, to a high-yielding japonica rice cultivar, Wuxiangjing 9, mediated with Agrobacterium tumeficien. And some selectable marker-free （SMF） transgenic lines had been subsequently selected. In present study, several homozygous transgenic lines with soft or waxy grain property were selected from the offspring of these SMF transformants containing the antisense Wx gene. The results from Northern blot analysis indicated that the levels of both the Wx mature mRNA and unspliced transcripts were reduced differently in the endosperm of different transgenic lines. The Wx protein level in mature seeds of the transgenic lines was also reduced differently. The amylose content of two transgenic lines was reduced to that of soft rice （about 10%） and one line was reduced to that of waxy rice （lower than 2%）. The gel consistency of these transgenic lines was softer than that of the wild type. The variance of gelatination temperature between the transgenic lines and the wild type did not reach to significant level. The most of the main agronomic traits of these transgenic lines were similar to that of the wild type. Therefore, three SMF transgenic homozygous lines with improved quality were successfully bred.