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Journal of Integrative Agriculture  2021, Vol. 20 Issue (7): 1880-1888    DOI: 10.1016/S2095-3119(21)63655-9
Special Issue: 昆虫合辑Plant Protection—Entomolgy 昆虫分子生物学合辑Insect Molecular Biology
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Functional analysis of the orphan genes Tssor-3 and Tssor-4 in male Plutella xylostella
LI Tian-pu1, 2, 3, ZHANG Li-wen1, 2, 3, LI Ya-qing1, 2, 3, YOU Min-sheng1, 2, 3, ZHAO Qian1, 2, 3 
1 State Key Laboratory of Ecological Pest Control for Fujian-Taiwan Crops, Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou 350002, P.R.China
2 Joint International Research Laboratory of Ecological Pest Control, Ministry of Education, Fuzhou 350002, P.R.China
3 Key Laboratory of Integrated Pest Management for Fujian-Taiwan Crops, Ministry of Agriculture and Rural Affairs, Fuzhou 350002, P.R.China
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孤儿基因是指在其他物种中没有同源序列的一类基因。在此,我们在小菜蛾中鉴定了两个孤儿基因,命名为Tssor-3Tssor-4。这两个基因都含有一个信号肽序列,表明它们具有分泌蛋白的功能。基于实时荧光定量PCR(qPCR)的表达模式分析表明,这两个孤儿基因均在除睾丸外的雄性生殖腺中特异表达;其表达量在雄成虫时期达到顶峰。免疫荧光实验表明,这两种蛋白均为精液蛋白,暗示它们在调节雄性生殖方面具有潜在的作用。为了进一步探索它们的功能,我们通过RNA干扰(RNAi)下调了这两个基因的表达量,结果表明,干扰后24 h,Tssor-3Tssor-4的表达量均显著低于对照组。生物测定实验表明,当Tssor-3Tssor-4基因表达量降低时,小菜蛾的产卵量和子代卵的孵化率均显著下降,表明这两个孤儿基因在小菜蛾雄性育性中起作用。我们的结果为孤儿基因参与雄性生殖调节提供了证据,这对雄性在进化过程中的适应性很重要。

Orphan genes are genes with no sequence homologues in other species.  Here, we identified two orphan genes, namely, Tssor-3 and Tssor-4, in Plutella xylostella.  Both genes contained a signal peptide sequence, suggesting their functions as secreted proteins.  Expression pattern analysis based on real-time quantitative PCR (qPCR) showed that both orphan genes were specifically expressed in all male gonads except the testes.  The expression of both the orphan genes peaked at the male adult stage.  Immunofluorescence assays suggested that the two proteins were seminal proteins, indicating their potential roles in male reproductive regulation.  To further explain their functions, we knocked down the expression of these two genes by RNA interference (RNAi).  The results showed that the expression of Tssor-3 and Tssor-4 was significantly downregulated at 24 h after injection compared to that of the controls.  Biological assays showed that the number of laid eggs and the hatching rate of offspring eggs were significantly reduced when the expression of Tssor-3 and Tssor-4 was reduced, suggesting that the two orphan genes played a role in male fertility in P. xylostella.  Our results provide evidence that orphan genes are involved in male reproductive regulation, which is important for male fitness during evolution.
Keywords:  Plutella xylostella        orphan genes        male fertility        RNA interference  
Received: 04 November 2020   Accepted: 02 June 2021
Fund: This work was supported by the Natural Science Foundation of Fujian Province, China (2020J01525), the National Natural Science Foundation of China (31320103922 and 31230061) and the Major Science and Technology Projects in Fujian Province, China (2018NZ010100130).
Corresponding Authors:  Correspondence ZHAO Qian, Tel: +86-591-83843724, E-mail:   
About author:  LI Tian-pu, E-mail:

Cite this article: 

LI Tian-pu, ZHANG Li-wen, LI Ya-qing, YOU Min-sheng, ZHAO Qian. 2021. Functional analysis of the orphan genes Tssor-3 and Tssor-4 in male Plutella xylostella. Journal of Integrative Agriculture, 20(7): 1880-1888.

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