糙皮侧耳Pleurotus ostreatus子实体凝集素基因polectin2及其启动子的克隆及功能鉴定

发布时间：2018-05-29 23:45

本文选题：糙皮侧耳 + 凝集素基因　；参考：《菌物学报》2017年08期

【摘要】：凝集素在食用菌的生长发育、抗病虫等逆境及抗肿瘤等方面发挥着重要的作用,但是关于糙皮侧耳凝集素基因的抗逆功能研究还比较薄弱。因此,本文利用PCR技术克隆得到糙皮侧耳子实体凝集素polectin2基因及其启动子序列,测序和生物信息学结果显示该基因不含内含子,完整开放阅读框为408bp,编码136个氨基酸,预测蛋白分子量15.3k Da;启动子polectinpro序列长度为927bp,生物数据库PLACE分析结果表明polectinpro含有低温、干旱胁迫诱导和病程相关等抗逆元件。然后,构建含有GFP融合蛋白的重组植物表达载体pCAMBIA1302-polectinpro-gfp,并进行烟草遗传转化,结果证明该启动子能够在低温诱导下驱动gfp基因表达。同时成功构建了含有polectin2的重组植物表达载体pCAMBIA1301-polectin2并进行了烟草遗传转化;对阳性转化植株T1代和野生型种子进行低温胁迫实验,结果表明转基因种子萌发率明显高于野生型。综上所述,推测polectin2基因具有耐低温功能,且其启动子具有低温诱导活性。上述结果为进一步利用子实体凝集素polectin2基因提高食用菌的耐低温等抗逆功能提供了理论依据和技术支持。
[Abstract]:Lectin plays an important role in the growth and development of edible fungi, resistance to diseases and insects, and anti-tumor, but the study on the anti-stress function of lectin gene of Pleurotus strigella is still weak. Therefore, the lectin polectin2 gene and its promoter sequence were cloned by PCR technique. Sequencing and bioinformatics showed that the gene contained no intron, and the complete open reading frame was 408 BP, encoding 136 amino acids. The molecular weight of predicted protein was 15.3kDa. the length of promoter polectinpro sequence was 927bp. the results of PLACE analysis in biological database showed that polectinpro contained anti-stress elements such as low temperature, drought stress induction and course of disease. Then, the recombinant plant expression vector pCAMBIA1302-polectinpro-gfpcontaining GFP fusion protein was constructed and transformed into tobacco. The results showed that the promoter could drive the expression of gfp gene under low temperature induction. At the same time, the recombinant plant expression vector pCAMBIA1301-polectin2 containing polectin2 was successfully constructed and genetic transformation of tobacco was carried out. The results showed that the germination rate of transgenic seeds was significantly higher than that of wild type. In conclusion, polectin2 gene has the function of low temperature tolerance and its promoter has low temperature induction activity. These results provide theoretical basis and technical support for the further use of the fruiting body lectin polectin2 gene to enhance the resistance to low temperature and other stresses of edible fungi.
【作者单位】：吉林农业大学农学院食药用菌教育部工程中心;龙泉市食药用菌产业办公室;
【基金】：公益性行业(农业)科研专项(201503137) 973项目(2014CB138305) 吉林省秸秆综合利用技术创新平台(吉高平合字[2014]B-1) 吉林农业大学校内启动基金(2015007) 吉林省科技发展项目(20140101148JC)~~
【分类号】：S646

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