褐飞虱热胁迫下内参基因的筛选及热激蛋白基因表达分析（英文）

【目的】褐飞虱Nilaparvata lugens(Stl)是为害水稻的重要害虫之一,温度是影响其暴发、迁飞的主要环境因子之一。本研究旨在探讨研究褐飞虱对高温胁迫适应性的热激蛋白基因表达调控模式。【方法】分别以不同的高温(30℃~40℃)处理褐飞虱雌、雄虫1 h和2 h,利用荧光定量PCR技术检测其体内的β-actin 1,β-actin2,β-actin3,28S rRNA,18S rRNA和α-2-tubulin 6个内参基因的表达量,用geN orm和BestK eeper软件分析确定最稳定表达的内参基因,并检测热胁迫后hsp70和hsp90基因在处理褐飞虱成虫体内的表达模式。【结果】geN orm软件分析结果表明,热胁迫后褐飞虱内参基因稳定性在雌虫体内为:β-actin1=β-actin328S rRNAα-2-tubulin18S rRNAβ-actin2;在雄虫体内为:β-actin1=β-actin3α-2-tubulin28S rRNA18S rRNAβ-actin2。BestK eeper软件分析结果显示,在热胁迫的雌、雄虫体内β-actin1均最稳定,18S rRNA次之,β-actin2最不稳定。两种软件分析结果基本一致。以β-actin1为校正内参基因,荧光定量PCR分析hsp70和hsp90在不同热胁迫条件下的表达模式,结果表明,各高温处理下hsp70表达量与对照26℃下的表达量没有显著性差异;而hsp90基因表达模式表现为被高温诱导上调表达,在雌、雄虫体内表达量达到最高的处理条件分别为40℃和38℃处理2 h。【结论】β-actin1基因可以作为热胁迫下褐飞虱雌雄虫体内基因表达模式分析的校正内参基因使用。褐飞虱hsp90基因能被高温诱导表达,该基因可能在褐飞虱适应热胁迫过程中起着重要的作用。

Identification of reference genes and expression analysis of heat shock protein genes in the brown planthopper,Nilaparvata lugens (Hemiptera: Delphacidae),after exposure to heat stress

【Aim】 The brown planthopper, Nilaparvata lugens (Stål) (Hemiptera: Delphacidae), is a serious rice pest in China and Southeast Asia. The occurrence and migration of N. lugens is thought to be related to temperature. This study was conducted to understand the expression patterns of heat shock protein genes (hsps) in the adaptation to temperature stress in N. lugens. 【Methods】 Female and male N. lugens adults were exposed to high temperature (30℃-40℃) for 1 h and 2 h, respectively. Real-time PCR was used to detect the expression of β-actin 1, β-actin 2, β-actin 3,28S rRNA, 18S rRNA and α-2-tubluin in their bodies. The most stable candidate reference gene was identified using geNorm and BestKeeper software. The expression levels of hsp70 and hsp90 genes in the treated N. lugensadults were measured using RT-qPCR. 【Results】 The most stable reference gene in both female and male adults of N. lugens after exposure to heat stress was β-actin 1. The expression levels of hsp70 after heat stress ranging from 30℃ to 40℃ in both female and male adults were not significantly different compared with those in the control group. The expression level of hsp 90 displayed significant up-regulation and reached the highest levels in female adults and male adults exposed to 40℃ and 38℃ for 2 h, respectively. 【Conclusions】β-actin 1 can be used as the reference gene for normalization of gene expression under high temperature stress in N. lugensadults. The expression of hsp 90 is induced by heat shock and the over-expression of hsp 90 might be involved in the enhancement of thermal tolerance in N. lugens adults.