›› 2014, Vol. 57 ›› Issue (6): 673-680.doi:

• 研究论文 • 上一篇    下一篇

桔小实蝇细胞凋亡基因hid的克隆及不同发育阶段表达分析

蔡玉音1,#, 武  强1,#, 刘桂清1,2, 吕志创1, 李建伟1, 张桂芬1, 万方浩1,*   

  1. (1. 中国农业科学院植物保护研究所, 植物病虫害生物学国家重点实验室, 北京 100193; 2. 广东省昆虫研究所, 广州 510260)
  • 出版日期:2014-06-20 发布日期:2014-06-20

Cloning and developmental expression analysis of an apoptosis gene hid in the oriental fruit fly, Bactrocera dorsalis (Diptera: Tephritidae)

AI Yu-Yin1, #, WU Qiang1, #, LIU Gui-Qing1,2, LÜ Zhi-Chuang1, LI Jian-Wei1, ZHANG Gui-Fen1, WAN Fang-Hao1,*   

  1. (1. State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China; 2. Guangdong Entomological Institute, Guangzhou 510260, China)
  • Online:2014-06-20 Published:2014-06-20

摘要: 【目的】细胞凋亡是一个细胞自动结束生命的过程,也称为程序性细胞死亡(programmed cell death, PCD)。头部退化缺陷基因 (head involution defective, hid )是昆虫细胞凋亡基因RHG家族的成员,该家族基因通过克服凋亡蛋白抑制因子(inhibitor of apoptosis proteins, IAPs)的保护作用来保证PCD的发生。本研究旨在克隆和分析桔小实蝇Bactrocera dorsalishid基因,并研究其在各发育阶段的表达情况。【方法】利用RT-PCR和RACE技术获得了桔小实蝇hid基因的cDNA全长序列。利用荧光定量PCR对hid基因在桔小实蝇不同发育阶段的转录水平进行分析。【结果】克隆获得桔小实蝇hid cDNA序列,将其命名为Bdhid (GenBank登录号为KJ461670),其开放阅读框长1 029 bp,编码342个氨基酸。氨基酸序列分析显示其具有一个短的N-端肽基元( IAP-binding-motif, IBM) 和C-端Grim Helix 3 (GH3) 结构域,与其他已知的双翅目昆虫hid基因有较高的同源性。实时荧光定量PCR检测结果表明,该基因在桔小实蝇的幼虫期表达水平较低,蛹期及成虫期表达量显著升高。【结论】这些结果为进一步研究hid基因在桔小实蝇细胞凋亡途径中的功能及其转基因条件致死品系的获得奠定了基础。

关键词: 桔小实蝇, 细胞凋亡基因, 分子克隆, 序列分析, 实时荧光定量PCR

Abstract: 【Aim】 Apoptosis is an active process that cells kill themselves automatically, and it is also referred to as programmed cell death (PCD). Head involution defective (hid) belongs to the RHG gene family whose members are primary regulators of PCD in insects due to their antagonistic effect on inhibitor of apoptosis proteins (IAPs).The aim of this study was to explore the expression pattern of hid in the oriental fruit fly, Bactrocera dorsalis at different developmental stages. 【Methods】 The full-length cDNA of hid was cloned by RT-PCR and RACE from B. dorsalis. The transcriptional level was analyzed by realtime quantitative PCR. 【Results】 The full-length cDNA sequence of hid from B. dorsalis was cloned, which was named as Bdhid (GenBank accession number: KJ461670), with a 1 029 bp open reading frame encoding 342 amino acids. It has the N-terminal IAP-binding-motif (IBM) and the C-terminal Grim Helix 3 motif (GH3) in its amino acid sequence, exhibiting high conservation with the known hid genes from other dipterans. The mRNA expression level of Bdhid was low in larvae and relatively higher in pupae and adults. 【Conclusion】 The results provide a foundation for further research of hid in insect apoptosis and for the development of conditional lethality strains by transgenic approaches.

Key words: Bactrocera dorsalis, apoptosis gene, molecular cloning, sequence analysis, real-time quantitative PCR