轉天山雪莲冷调节蛋白基因siCOR烟草的抗寒性鉴定.docVIP

转天山雪莲冷调节蛋白基因siCOR烟草的抗寒性鉴定 郭新勇，程晨，喻娜，祝建波（石河子大学 生命科学学院，石河子大学农业生物技术重点实验室，石河子 832003） 摘要：cDNA文库中随机挑取单克隆，采用通用引物M13进行PCR扩增出基因序列并测序，然后将该序列在GenBank中进行比对，根据该序列设计特异性引物进行PCR，最终获得了冷调节蛋白基因siCOR的完整开放读码框（ORF）PCR和RT-PCR对转化烟草进行鉴定，收取种子后筛选出纯合的T3株系，最后通过冷冻胁迫进行抗寒性分析。结果表明，siCOR大小661bp，与禾本科植物粳稻（Oryza sativa Japonica Group）的冷调节蛋白基因相似性最高，为65.04%；与野生型植株相比，转siCOR基因烟草均长有浓绿的叶片，无分支侧芽，生长快；在冷冻胁迫条件下，转siCOR基因的烟草叶片相对含水量、PSⅡ相对量子产率降低幅度、相对电导率和丙二醛含量的升高幅度均低于野生型烟草植株，而且转siCOR基因烟草植株叶片出现萎蔫症状的时间较迟、程度较轻，受到的伤害也较轻。可见，在冷冻胁迫条件下，转siCOR基因烟草植株表现出良好的生理和生长优势，显示出了较强的抗寒特征。 关键词： 天山雪莲；siCOR基因；烟草；抗寒性Q789 文献标识码：A Cold Resistance Identification of Tobacco Plants Transformed by siCOR of Sasussured involucrata Kar.et Kir GUO Xin-yong Cheng Chen YU Na ZHU Jian-bo ( Laboratory of Agricultural Biotechnology, College of life science of Shihezi University, Shihezi Xinjiang , 832003,China ） Abstract：comparing the sequence in GenBank, designing specific primer according to the sequence, finally we obtained the whole ORF of siCOR.Then plant expression vector pBIN438-siCOR was constructed and NC89 tobacco plants were transformed via Agrobacterium tumefaciens, PCR and RT-PCR analysis indicated that siCOR had been integrated into the genome. Homozygous transgenic tobacco plants were used in cold resistance identification. The results show that, the size of siCOR cDNA was 661bp, the identity of amino acid sequence was about 65.04% with Oryza sativa Japonica Group. siCOR tobacco has deep green leaves, grows fast and has no lateral bud compared with wild type; under the cold stress, the decrease of relative water content, reduction of relative quantum yield of PSⅡ, increase of relative conductivity and MDA of siCOR tobacco are lesser than control tobacco evidently, besides, transgenic plants has slow phenotype and lighter symptom. According to the above, we can draw that, under the cold stress, siCOR tobacco shows well growth traits and strong cold resistance characteristics. Key words：Sasussured involucrata Kar.et Kir；siCOR ge