云南药用野生稻高光效基因的筛选及功能分析

Breeding high photosynthetic efficiency of rice is the way to improve the yield and solve huge demand for food. Wild rice Oryza Officinalis Wall. evolved of some excellent traits during long-term of natural selection such as a strong growth potential with more than 20 times that of cultivated varieties, a great accumulation of biomass, and a higher net photosynthetic rate. And our result also indicated that the carboxylation efficiency and light saturation point of O.Officinalis of Yunnan was 2 times more than that of cultivated varieties. Our past research, a genomic BIBAC library of O. Officinalis of Yunnan was been constructed and used to transform Nipponbare for introgression lines. And among of the introgression lines, some T1 generations with higher net photosynthetic rate had been identified. This study, based on the T2 generations of introgression lines with high photosynthetic efficiency trait that screened out by measuring saveral biological characters, is to predict high photosynthetic efficiency candidate genes from T2 generations using genomic resequencing and RNA-Seq technology, that is, genomic resequencing can find out the differential DNA sequences from O. Officinalis when compared with Nipponbare genome, and then according to these DNA sequences, probes are designed to screen and sequence BIBAC clones to obtain the intact target DNA sequences; RNA-Seq technology can find out the differentially expressed genes between T2 generations of introgression lines and Nipponbare, witch is helpful to screen candidate genes from the target DNA. Ultimately, after transformating candidate genes into introgression lines and Nipponbare to identify their functions by RNAi and overexpression technics rerepectively, the high photosynthetic efficiency genes of O. Officinalis of Yunnan are determined. This will, to some extent, reveal the genetic basis of strong growth potential and great biomass accumulation traits of O. Officinalis, find out genes that can be applied to improve the photosynthesis and yield potential of cultivated varieties, which is important theoretical significance and application value.

培育高光效水稻是提高水稻产量和解决粮食需求的有效途径，高光效基因发掘在其中尤为重要。药用野生稻(O. Officinalis)具备高光效特性：强生长势、20倍以上于栽培稻的生物量积累和较高的净光合速率。我们发现云南药用野生稻的羧化效率、光饱和点均为栽培稻的2倍以上。前期，我们构建了云南药用野生稻基因组BIBAC文库，转入栽培稻日本晴建立了渗入系库，获得了净光合速率高于日本晴的渗入系T1代株系。本项目拟加代T1代株系到T2代，测定T2代株系的生物学性状，筛选出高光效T2代株系；利用基因组重测序和RNA-Seq技术分析渗入系T2代株系中来自云南药用野生稻基因组的DNA和基因表达特征，结合筛选BIBAC克隆测序，预测候选基因；通过基因沉默和过表达途径验证和分析候选基因功能，从中鉴定出云南药用野生稻高光效基因。这不仅对揭示云南药用野生稻高光效机理有科学意义，而且在培育高光效水稻材料方面有实践意义。

项目旨通过构建药用野生稻渗入系，对渗入系材料进行光合作用鉴定筛选出高光效株系，并从中筛选出药用野生稻后代的光合作用相关基因。.通过项目实施，对药用野生稻的光合作用水平进行了综合评价，明确了药用野生稻具有高羧化效率和光饱和点高的特点，同时在叶面积上有较大的优势，并进行了叶绿体超微结构的分析，综合确定了药用野生稻的高光效特性；利用野栽远缘杂交，克服种间生殖隔离构建了药用野生稻渗入系材料600余个株系，并对药用野生稻渗入系材料进行光合作用鉴定，筛选到7份高光效渗入系材料；利用药用野生稻转录组数据，对药用野生稻后代的光合作用相关基因进行分析，筛选出药用野生稻所特有的4个光合途径相关基因：PEPcase、Rubis-subs-bind、rbc-S、PPDK。并对这些基因进行了了表达和同源性分析，最终筛选到2个药用野生稻后代的C4途径PEPC和PPDK的同源基因；通过对筛选出的典型高光效渗入系材料和低光效渗入系材料进行转录组和蛋白组分析，从中初步筛选出27个光合作用相关差异表达基因，13个光合作用相关差异表达蛋白，并且初步明确了差异基因和蛋白在光合作用中的代谢调控通路。.项目构建的药用野生稻渗入系材料、筛选出的药用野生稻后代的光合相关基因，将为后续基因功能的深入研究和高产育种利用打下基础，具有科学意义和潜在应用价值。.项目发表研究论文5篇，其中SCI 1篇，申请发明专利3项，已授权1项。参与出版专著1部，培养硕士研究生1名。