意大利蝗海藻糖酶基因克隆及卵低温驯化下的表达模式

doi:10.16380/j.kcxb.2022.02.004

昆虫学报 ›› 2022, Vol. 65 ›› Issue (2): 157-166.doi: 10.16380/j.kcxb.2022.02.004

意大利蝗海藻糖酶基因克隆及卵低温驯化下的表达模式

刘倩¹, 罗迪¹, Roman JASHENKO², 季荣^1,*

(1. 新疆师范大学生命科学学院, 中亚区域跨境有害生物联合控制国际研究中心, 新疆特殊环境物种保护与调控生物学自治区重点实验室, 新疆特殊环境物种多样性应用与调控教育厅重点实验室, 乌鲁木齐 830054; 2. 哈萨克斯坦阿勒法拉比国立大学, 阿拉木图 050038)

出版日期:2022-02-20 发布日期:2022-03-10

Cloning of trehalase genes in Calliptamus italicus (Orthoptera: Acrididae) and their expression patterns in eggs under cold acclimation

LIU Qian¹, LUO Di¹, Roman JASHENKO², JI Rong^1,*

(1. International Research Center for the Collaborative Containment of CrossBorder Pests in Central Asia, Xinjiang Key Laboratory of Special Species Conservation and Regulatory Biology, Key Laboratory of Special Environment Biodiversity Application and Regulation in Xinjiang, Xinjiang Normal University, Urumqi 830054, China; 2. Al-Farabi Kazakh National University, Almaty 050038, Kazakhstan)

Online:2022-02-20 Published:2022-03-10

摘要/Abstract

摘要： 【目的】本研究通过克隆意大利蝗Calliptamus italicus卵海藻糖酶基因，分析其在蝗卵不同发育阶段的表达及低温驯化后的表达变化，阐明海藻糖酶基因在意大利蝗卵抵御低温胁迫过程中的作用。【方法】利用RT-PCR技术扩增蝗卵海藻糖酶基因的ORF全长序列，并进行生物信息学分析；利用RT-qPCR分析检测常温(27℃)和低温(0℃)驯化15 d海藻糖酶基因在蝗卵早期、滞育和滞育解除阶段的表达变化。【结果】克隆获得意大利蝗卵4个海藻糖酶基因，分别命名为CiTreM1, CiTreM2， CiTreS1和CiTreS2(GenBank 登录号分别为MZ669810, MZ669811, MZ669812和MZ669813)。其编码蛋白中，CiTreM1和CiTreM2为膜结合型海藻糖酶，CiTreS1和CiTreS2为可溶型海藻糖酶。CiTre基因在意大利蝗卵各发育阶段均表达，常温(27℃)下，CiTreS1和CiTreS2在卵早期阶段表达水平最高，CiTreM2在卵滞育阶段表达水平最高，CiTreM1在卵滞育解除阶段表达水平最高。低温(0℃)驯化15 d后CiTreM1和CiTreM2表达量在意大利蝗卵发育的3个阶段均增加，CiTreS1和CiTreS2仅在蝗卵早期阶段表达量下降，其他阶段表达量增加；低温驯化后CiTreM1和CiTreM2的表达量在意大利蝗卵早期阶段增幅最大，CiTreS1和CiTreS2在蝗卵滞育解除阶段的增幅最大。低温(0℃)驯化15 d后，在意大利蝗卵同一阶段4个CiTre基因的表达量明显不同，卵早期阶段和滞育阶段CiTreM2表达量最大，滞育解除阶段CiTreS1表达量最大。【结论】克隆获得的4个意大利蝗卵海藻糖酶基因均参与了蝗卵发育过程，根据低温驯化前后表达量变化幅度判断，CiTreM2和CiTreS1在意大利蝗卵抵御低温胁迫过程中发挥着关键作用。

关键词: 意大利蝗, 胚胎发育, 海藻糖酶基因, 低温驯化, 表达谱

Abstract: 【Aim】This study aims to clarify the role of trehalase genes in the resistance to low temperature stress in Calliptamus italicus eggs by cloning trehalase genes in C. italicus eggs, and analyzing their expression profiles in eggs at different developmental stages and the expression changes after cold acclimation. 【Methods】The full-length ORF sequences of trehalase genes from C. italicus eggs were amplified by RT-PCR, and analyzed by bioinformatics tools. RT-qPCR was used to detect the expression changes of trehalase genes in C. italicus eggs at the early stage, diapause stage and diapause termination stage under the room temperature (27℃) and after acclimation to low temperature (0℃) for 15 d. 【Results】Four trehalase genes from C. italicus eggs were cloned and named CiTreM1, CiTreM2, CiTreS1 and CiTreS2, with the GenBank accession numbers of MZ669810, MZ669811, MZ669812 and MZ669813, respectively. Among their encoded proteins, CiTreM1 and CiTreM2 are membrane-bound trehalases, and CiTreS1 and CiTreS2 are soluble trehalases. CiTre genes were expressed in C. italicus eggs at various developmental stages. At the room temperature (27℃), CiTreS1 and CiTreS2 had the highest expression level in C. italicus eggs at the early stage, while CiTreM2 and CiTreM1 had the highest expression level in eggs at the diapause stage and diapause termination stage, respectively. After acclimation to low temperature (0℃) for 15 d， the expression levels of CiTreM1 and CiTreM2 increased in eggs at all the three stages, while those of CiTreS1 and CiTreS2 decreased only in eggs at the early stage but increased in eggs at the other stages. After cold acclimation, the expression levels of CiTreM1 and CiTreM2 increased the most in C. italicus eggs at the early stage, while those of CiTreS1 and CiTreS2 increased the most in eggs at the diapause termination stage. After acclimation to low temperature (0℃) for 15 d, the expression levels of the four CiTre genes were obviously different in C. italicus eggs at the same stage, and CiTreM2 showed the highest expression level in C. italicus eggs at early stage and the diapause stage, and CiTreS1 showed the highest expression level in eggs at the diapause termination stage. 【Conclusion】The four cloned trehalase genes in C. italicus eggs are all involved in the development of C. italicus eggs. According to the changes in expression level before and after cold acclimation, CiTreM2 and CiTreS1 play a key role in the process of C. italicus eggs resisting low temperature stress.

Key words: Calliptamus italicus, embryonic development, trehalase gene, cold acclimation, expression profile

刘倩, 罗迪, Roman JASHENKO, 季荣. 意大利蝗海藻糖酶基因克隆及卵低温驯化下的表达模式[J]. 昆虫学报, 2022, 65(2): 157-166.

LIU Qian, LUO Di, Roman JASHENKO, JI Rong. Cloning of trehalase genes in Calliptamus italicus (Orthoptera: Acrididae) and their expression patterns in eggs under cold acclimation[J]. Acta Entomologica Sinica, 2022, 65(2): 157-166.

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意大利蝗海藻糖酶基因克隆及卵低温驯化下的表达模式

Cloning of trehalase genes in Calliptamus italicus (Orthoptera: Acrididae) and their expression patterns in eggs under cold acclimation

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