昆虫学报 ›› 2023, Vol. 66 ›› Issue (12): 1560-1569.doi: 10.16380/j.kcxb.2023.12.003

• 研究论文 • 上一篇    下一篇

柞蚕微孢子虫感染后柞蚕卵转录组及免疫相关基因功能分析

徐欣1,2, 吴玉娇1, 于滨1, 孟宪志1, 陈杰1, 刘中文2, 张永君2, 潘国庆1,*   

  1. (1.西南大学, 微孢子虫感染与防控重庆市重点实验室, 重庆 400715; 2. 河南省蚕业科学研究院, 郑州 450008)
  • 出版日期:2023-12-20 发布日期:2024-01-21

Transcriptome and immune-related gene function analyses of Antheraea pernyi (Lepidoptera: Saturniidae) eggs infected by Nosema pernyi

XU Xin1,2, WU Yu-Jiao1, YU Bin1, MENG Xian-Zhi1, CHEN Jie1, LIU Zhong-Wen2, ZHANG Yong-Jun2, PAN Guo-Qing1,*   

  1.  (1. Chongqing Key Laboratory of Microsporidia Infection and Control, Southwest University, Chongqing 400715, China; 2. Sericultural Research Institute of Henan Province, Zhengzhou 450008, China)
  • Online:2023-12-20 Published:2024-01-21

摘要: 【目的】 柞蚕微孢子虫Nosema pernyi引发柞蚕Antheraea pernyi微粒子病,威胁柞蚕种质资源的保育及生产安全,并且能够经卵垂直传播,因此需要寻找柞蚕体内应对柞蚕微孢子虫侵染后的免疫基因,为后续寻找和培育抗微粒子病新品种提供帮助。【方法】 构建柞蚕微孢子虫侵染后柞蚕卵转录组,进行GO功能注释和KEGG分析确定免疫相关基因;PCR克隆柞蚕谷胱甘肽S-转移酶(glutathione S-transferase, GST)基因ApGSTo1并进行生物信息学分析;运用RT-qPCR检测ApGSTo1在柞蚕微孢子虫侵染柞蚕产出后0-10 d卵中的表达量。【结果】 在感染与未感染柞蚕微孢子虫柞蚕卵转录组检测到的表达基因数为17 453,其中显著差异表达基因数目为89个; GO功能注释中细胞解剖实体和催化活性相关基因数量最多,KEGG分析中参与运输和分解代谢、信号转导及消化系统这3个通路的基因数量最多。基于柞蚕微孢子虫侵染后柞蚕卵转录组数据分析,将ApGSTo1作为目的基因,PCR克隆及系统进化分析结果证明ApGSTo1属于omega家族GST;感染柞蚕微孢子虫柞蚕卵中ApGSTo1表达量比正常对照组的高,且在产卵后2 d时的表达量极显著高于正常对照组的。【结论】 获得了柞蚕微孢子虫侵染后柞蚕卵转录组数据,找到了1个免疫相关基因ApGSTo1,从分子生物学和转录水平初步证明了ApGSTo1参与柞蚕卵内的抗柞蚕微孢子虫侵染,还发现了抗柞蚕微孢子虫侵染反应的关键时间为产卵后2 d时,这些结果为后续研究ApGSTo1在抗逆性和氧化还原等功能方面提供研究基础。

关键词: 柞蚕, 柞蚕微孢子虫, GST, ApGSTo1, 抗逆性

Abstract: 【Aim】 Nosema pernyi can cause the tussah microparticle disease of Antheraea pernyi, threatening the conservation and production safety of A. pernyi germplasm resources, and be vertically transmitted through eggs. Therefore, it is necessary to search for immune genes in A. pernyi to deal with the N. pernyi infection, so as to provide assistance for the subsequent search and cultivation of new varieties resistant to tussah microparticle disease. 【Methods】 The transcriptome of A. pernyi eggs infected with N. pernyi was constructed, and the immune-related genes were identified through GO functional annotation and KEGG analysis. The glutathione S-transferase (GST) gene of A. pernyi ApGSTo1 was cloned by PCR and bioinformatically analyzed. RT-qPCR was used to detect the expression level of ApGSTo1 in the 0-10-day-old eggs of A. pernyi infected by N. pernyi.【Results】 The number of expressed genes detected in the transcriptome of A. pernyi eggs infected and uninfected with N. pernyi was 17 453, and we found 89 significantly differentially expressed genes. In GO functional annotation, the numbers of genes involved in cellular anatomical entity and catalytic activity were the highest, while in KEGG analysis, the numbers of genes involved in the three pathways of transportation and catabolism, signal transduction, and digestive system were the highest. Based on transcriptome data analysis of A. pernyi eggs infected by N. pernyi, ApGSTo1 was used as the target gene. Results of PCR cloning and phylogenetic analysis proved that ApGSTo1 belongs to the omega-class GST. The expression level of ApGSTo1 in the A. pernyi eggs infected by N. pernyi was higher than that of the normal control group, and the expression level of ApGSTo1 in the 2-day-old eggs of A. pernyi infected by N. pernyi was extremely significantly higher than that of the normal control group. 【Conclusion】 Transcriptome data of A. pernyi eggs infected by N. pernyi were obtained, and an immune-related gene ApGSTo1 was identified. It was preliminarily demonstrated from molecular biology and transcriptional levels that ApGSTo1 participates in the stress resistance to N. pernyi infection in A. pernyi eggs. It was also found that the key time for the stress resistance to N. pernyi infection in A. pernyi eggs is at 2 d after egg laying. These results provide a research foundation for future research on the functions of ApGSTo1 in stress resistance and redox.

Key words: Antheraea pernyi, Nosema pernyi, GST, ApGSTo1, resistance