褐飞虱两个酚氧化酶原基因的特性及细菌诱导后的表达分析

doi:10.16409/j.cnki.2095-039x.2018.01.004

中国生物防治学报 ›› 2018, Vol. 34 ›› Issue (1): 44-51.DOI: 10.16409/j.cnki.2095-039x.2018.01.004

褐飞虱两个酚氧化酶原基因的特性及细菌诱导后的表达分析

陈静¹, 张道伟², 钱正敏²

1. 遵义医学院, 遵义 563000;
2. 遵义师范学院生命科学学院/赤水河流域动物资源保护与应用研究重点实验室, 遵义 563002

收稿日期:2017-06-23 出版日期:2018-02-08 发布日期:2018-02-06
通讯作者: 张道伟,教授,E-mail:zhangdw1000@163.com
作者简介:陈静,博士,副教授,E-mail:chenjing_1983@126.com
基金资助:
国家自然科学基金（31560511）；贵州省科学技术基金（2014-2014）；贵州省教育厅青年基金（2014-320）

Characteristics and Bacterial-induced Expression Analysis of Two Prophenoloxidase Genes in The Brown Planthopper, Nilaparvata lugens (Hemiptera Delphacidae)

CHEN Jing¹, ZHANG Daowei², QIAN Zhengmin²

1. Zunyi Medical College, Zunyi 563000, China;
2. Key Laboratory of Protection and Utilization of Animal Resource in Chishui River Basin/College of Biotechnology and Agriculture, Zunyi Normal College, Zunyi 563002, China

Received:2017-06-23 Online:2018-02-08 Published:2018-02-06

摘要/Abstract

摘要： 为探讨褐飞虱Nilaparvata lugens（Stål）酚氧化酶原基因的发育表达模式及生物学功能，利用转录组数据和RACE方法获得两条褐飞虱酚氧化酶原基因NlPPO1（GenBank No.：ALE32751）和NlPPO2（GenBank No.：ALE32752）全长序列，褐飞虱NlPPO1全长为2316 bp，CDS编码区为2073 bp，编码690氨基酸；NlPPO2全长为2738 bp，CDS编码区为2100 bp，编码699氨基酸。通过实时荧光定量PCR方法检测NlPPO的发育表达模式及注射细菌诱导后NlPPO的表达量变化。发育表达模式结果显示，NlPPO1和NlPPO2在肠中都有较高的表达水平，而NlPPO1在表皮细胞表达量最低，NlPPO2在脂肪体表达量最低；在5龄幼虫阶段NlPPO1随时间增加表达量增加，NlPPO2表达量则维持相对较高水平；在成虫阶段NlPPO1表达量维持在较低水平，NlPPO2随时间增加表达量降低。注射大肠杆菌诱导NlPPO结果表明，诱导24 h后，NlPPO1表达量显著升高，NlPPO2表达量变化差异不显著；注射枯草芽胞杆菌诱导NlPPO结果表明，诱导24 h后，NlPPO1和NlPPO2表达量都显著升高。研究结果显示褐飞虱NlPPO1和NlPPO2的发育表达模式存在差异，对不同菌诱导免疫应答也存在差异，暗示二者具有不同的生物学功能。该结果为进一步探索酚氧化酶原在褐飞虱对病原菌的免疫响应机制奠定基础。

关键词: 褐飞虱, 酚氧化酶原, 特性, 表达模式, 诱导表达

Abstract: This study aims to explore the developmental expression pattern and biological function of prophenoloxidase in Nilaparvata lugens. The cDNA sequences encoding NlPPO1 (GenBank accession number:ALE32751) and NlPPO2 (GenBank accession number:ALE32752) were obtained from RNA-seq transcriptome sequencing database and rapid amplification of cDNA ends (RACE). The sequence analysis showed that the full-length cDNA sequence of NlPPO1 is 2316 bp, which contains an open reading frame (ORF) of 2073 bp encoding 690 amino acid-residues. And the full-length cDNA sequence of NlPPO2 is 2738 bp, which contains an ORF of 2100 bp encoding 699 amino acid-residues. The expression patterns and the changes of NlPPO mRNA induced by injecting bacteria were analyzed by real-time quantitative PCR. The developmental expression patterns showed that NlPPO1 and NlPPO2 were highly expressed in the gut, with a low mRNA level in the epidermis cell and fat body, respectively. The NlPPO1 mRNA expression increased with time and the NlPPO2 mRNA expression remained at a relatively high level in 5th instar larvae stage. The NlPPO2 mRNA expression decreased with time and the NlPPO1 mRNA expression remained at a relatively low level in adult stage. The NlPPO1 mRNA expression level significantly increased while the NlPPO2 mRNA expression level did not change significantly at 24 h after injection of Escherichia coli in N. lugens. The NlPPO1 and NlPPO2 mRNA expression levels significantly increased at 24 h after injection of Bacillus subtilis in N. lugens. The results showed that there are some differences between NlPPO1 and NlPPO2 between developmental stages and tissues in N. lugens. And the mRNA expression levels of NlPPO1 and NlPPO2 differ in the immune responses to bacteria, suggesting that the two NlPPO genes have different biological functions. The results provide a good foundation for further research on immune responses of NlPPO in the brown planthopper.

Key words: Nilaparvata lugens, prophenoloxidase, characteristics, expression pattern, inducible expression

中图分类号:

Q966

陈静, 张道伟, 钱正敏. 褐飞虱两个酚氧化酶原基因的特性及细菌诱导后的表达分析[J]. 中国生物防治学报, 2018, 34(1): 44-51.

CHEN Jing, ZHANG Daowei, QIAN Zhengmin. Characteristics and Bacterial-induced Expression Analysis of Two Prophenoloxidase Genes in The Brown Planthopper, Nilaparvata lugens (Hemiptera Delphacidae)[J]. journal1, 2018, 34(1): 44-51.

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褐飞虱两个酚氧化酶原基因的特性及细菌诱导后的表达分析

Characteristics and Bacterial-induced Expression Analysis of Two Prophenoloxidase Genes in The Brown Planthopper, Nilaparvata lugens (Hemiptera Delphacidae)

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