光周期和温度对生物钟基因cwo在 棉铃虫幼虫节律表达的影响
The effect of photoperiod and temperature on the diurnal expression of the circadian clock gene cwo in larvae of cotton bollworm, Helicoverpa armigera (Hübner)
刘孝明1** 张松斗1 马木提·赛丽蔓2 邹 驰1 李 贞1 张青文1 刘小侠1*
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DOI：10.7679/j.issn.2095-1353.2016.117
作者单位：1中国农业大学昆虫系，北京 100193；2. 新疆博尔塔拉蒙古自治州农业技术推广中心，博乐 83340
中文关键词：棉铃虫幼虫，生物钟基因，光周期，温度，昼夜节律，转录表达
英文关键词：Helicoverpa armigera larvae, circadian clock gene, photoperiod, temperature, circadian rhythm, transcript expression
中文摘要：
【目的】 生物钟普遍存在于生物体的生长发育、行为及生理等各个方面。本研究旨在探究光周期和温度对生物钟基因cwo在棉铃虫Helicoverpa armigera幼虫昼夜节律表达的影响。【方法】 通过转录组测序获得了生物钟基因cwo的cDNA开放阅读框全长，利用实时荧光定量PCR技术分析了生物钟基因cwo在棉铃虫幼虫中的时空表达，以及其在不同光周期和不同温度条件下的昼夜表达节律。【结果】 序列分析结果显示，棉铃虫生物钟基因cwo的cDNA序列开放阅读框为1 335 bp，编码444个氨基酸残基，预测蛋白的分子质量为49.95 ku。时空表达分析表明，cwo基因在棉铃虫幼虫组织中以脑和腹神经索表达量较高，在卵孵化期、蜕皮期、蛹变态期及羽化前1 d高峰度表达。头部昼夜节律表达分析表明，正常光周期条件下生物钟基因cwo呈昼夜节律表达，在暗期高表达；短时间持续暗期/光期处理使cwo表达的时间相位提前，长时间持续暗期/光期处理使cwo表达的节律性显著降低；38℃处理使cwo表达短期内显著上调，并改变了其节律性，18℃处理使cwo表达在暗期显著下调，但是节律性不变。【结论】 生物钟基因cwo在棉铃虫幼虫头部的昼夜节律表达受到光周期和温度的影响。
英文摘要：
[Objectives] Circadian clock is easily traced in the growth, behavior, physiology and other aspects of organisms. To explore the effect of photoperiod and temperature on the diurnal expression of the circadian clock gene cwo in cotton bollworm, Helicoverpa armigera, larvae. [Methods] The full-length ORF (open reading frame) of cwo cDNA was obtained through transcriptome sequencing. The spatial and temporal expression of cwo and its circadian rhythm of expression under different photoperiod and temperature conditions were analyzed with real-time PCR. [Results] Sequence analysis showed that the ORF of the cotton bollworm cwo cDNA sequence was 1 335 bp, encoding 444 amino acid residues, and had a predicted protein molecular mass of 49.95 ku. Spatial and temporal expression analysis showed that the cwo gene was more highly expressed in the brain and ventral nerve cord than in other tissues. Transcription of this gene peaked during the hatching stage, the molting period, larval-pupal metamorphosis and 1 d before eclosion. Analysis of the expression of this gene in the head of cotton bollworm larvae indicated that under normal photoperiod conditions the expression of cwo had an obvious circadian rhythm and peaked during the scotophase. Under a short scotophase, or photophase, the timing of cwo expression was advanced, whereas long scotophase or photophase significantly decreased the circadian oscillation of cwo expression. cwo expression was significantly up-regulated in the short term at 38℃, and its rhythm was disturbed, whereas at 18℃ expression was significantly down-regulated during the scotophase and its rhythm was unchanged. [Conclusion] The circadian expression of the circadian clock gene cwo in larvae of the cotton bollworm is influenced by photoperiod and temperature.