韭菜迟眼蕈蚊紫外敏感视蛋白基因的克隆及光强度对其表达量影响

为明确韭菜迟眼蕈蚊Bradysia odoriphaga紫外敏感视蛋白基因Bo-uv的作用及其与趋光性的关系,利用常规PCR方法克隆获得Bo-uv基因的全长cDNA序列,分析了其敏感视蛋白的氨基酸序列与其它12种昆虫同源蛋白氨基酸序列之间的系统进化关系,运用qPCR技术检测了不同发育阶段、不同组织及不同光强度下Bo-uv基因的相对表达量。结果表明,Bo-uv基因cDNA全长2 757 bp,开放阅读框1 542 bp,编码514个氨基酸。韭菜迟眼蕈蚊紫外敏感视蛋白的氨基酸序列与其它12种昆虫同源蛋白的氨基酸序列一致性为21.93%～43.00%,与橘小实蝇Bactrocera dorsalis的氨基酸序列同源性最高。Bo-uv基因在韭菜迟眼蕈蚊蛹末期、成虫期表达,在成虫头部的相对表达量较高。在0～10 000 lx光强范围内雌、雄成虫体内该基因的相对表达量均呈先增高后降低趋势。与对照相比,1 000 lx光强度下其相对表达量显著升高,10 000 lx时相对表达量显著降低。表明光强度能够有效地调控Bo-uv基因的表达,该基因在韭菜迟眼蕈蚊感知外界光刺激过程中具有重要作用。

Molecular cloning of an ultraviolet wavelength-sensitive opsin gene in Chinese chive gnat Bradysia odoriphaga and gene expression in different light intensities

To interpret the functions of an ultraviolet wavelength-sensitive gene (Bo-uv) and the relationship between Bo-uv gene and phototaxis in Chinese chive gnat Bradysia odoriphaga, the fulllength cDNA of Bo-uv was cloned by PCR, and the phylogenetic relationships between the amino acid sequences of its opsin and those of other 12 insects''homologous proteins were analyzed. The relative expression levels of the gene in different developmental stages and different tissues of adults were determined under four different light intensities by qPCR. The results showed that the full-length cDNA of the Bo-uv gene was 2 757 bp, which contained a 1 542 bp ORF encoding 514 amino acids, and the amino acid sequence of ultraviolet wavelength-sensitive opsin in Bradysia odoriphaga shared an identity of 21.93%-43.00% with those of 12 homologous proteins, with the highest homology with Bactrocera dorsalis. The Bo-uv gene was mainly expressed by the end of pupal stages and adults, and the relative expression level in the head was most highly expressed. The results showed that the expression of Bo-uv opsin gene firstly decreased, and then increased with increasing light intensity in the female and male adults. Compared with the control, the relative expression level of Bo-uv gene significantly decreased under 1 000 lx and the relative expression level of this gene significantly increased under 10 000 lx. The expression level of Bo-uv gene was controlled effectively by light intensity, indicating that Bo-uv gene was very important for perceiving ambient light stimuli.