异色瓢虫多巴脱羧酶基因序列及低温诱导表达分析  

Gene sequence and expression analysis of Dopa decarboxylase in Harmonia axyridis under low temperature induction

作　　者：汪慧娟[1] 施佐堃 徐风娇 沈祺达[1] 徐燕霞[1] 王世贵[1] 唐斌[1] 

机构地区：[1]杭州师范大学,动物适应与进化杭州市重点实验室,杭州310036

出　　处：《环境昆虫学报》2016年第2期271-279,共9页Journal of Environmental Entomology

基　　金：国家自然科学基金(31371996);杭州市科技局计划项目(20140432B01);杭州师范大学优秀中青年教师支持计划(JTAS2011-01-031)

摘　　要：多巴脱羧酶(dopa decarboxylase,DDC)是把多巴降解成多巴胺的重要酶类,在昆虫行为和发育中具有重要作用。本研究在转录组获得异色瓢虫Harmonia axyridis DDC基因序列的基础上,从ORF两端设计特异性引物进行扩增并测序验证,获得了DDC基因的c DNA的ORF全长序列,其包含1431 bp,编码476个氨基酸,软件分析预测显示该基因编码蛋白的分子量为53.88 k Da,理论等电点为5.80。本实验分为升温、降温、低温储存以及不同发育阶段(包括预蛹,1-3 d蛹,1-3 d 4龄幼虫以及羽化1-3 d成虫)4个处理组,采用荧光定量PCR技术研究异色瓢虫不同处理组DDC基因的表达水平。结果表明:DDC基因在蛹期第1天的表达量最高,在升温诱导条件下表达无差异,降温诱导下表达量上升;黄色雌成虫低温储存条件下基因表达量先显著上升后显著下降,黑色雌成虫表达量无差异,表明DDC基因可以在低温胁迫下高表达促使异色瓢虫适应环境变化。Dopa decarboxylase（ DDC） is an important enzyme,which can degrade levodopa into dopamine and play key role in the insect behavior and develpoment. Based on Harmonia axyridis transcriptome,the full c DNA of DDC in H. axyridis have been found and its accuracy identified by PCR amplification and sequenced. The whole DDC gene consists of 1431 bp open reading frame（ OFR） encoding 476 amino acids by PCR and the specific primers from ORF at both ends of the design. Software analysis predicts DDC has a molecular mass of 53. 88 k Da and isoelectric point of 5. 80. Meantime real-time PCR was adopted to analyze the expression of H. axyridis DDC at four treatment groups,which including different developmental stages from 4thinstar larva to adults,under warming and cooling treatments,and in the low temperature storage treatment. The results show that the expression level of DDC is highest in the first day of pupal period,and the expression level of DDC gene is indifference under warming process but up-regulation under cooling process. DDC gene expression of yellow female individuals is significantly up-regulated then down-regulated but the expression of black female individuals is indifference under the treatment of low temperature storage.The results imply that DDC gene can help H. axyridis adapt to environment changes by precisely theregulation of the dopa decarboxylase synthesis.

关 键 词：多巴脱羧酶 异色瓢虫 实时荧光定量PCR 低温诱导 表达分析 

分 类 号：Q963[生物学—昆虫学] S476[农业科学—农业昆虫与害虫防治]