<i>CYP</i>307A1 RNAi重组小球藻口服喂饲对白纹伊蚊的致死作用

黄小丹, 肖洒, 贺长皓, 李智杰, 张秀霞, 李亚军, 费小雯, 邓晓东

doi:10.11853/j.issn.1003.8280.2023.03.010

中国媒介生物学及控制杂志 >

2023 , Vol. 34 >Issue 3: 336 - 343

DOI: https://doi.org/10.11853/j.issn.1003.8280.2023.03.010

实验研究

CYP307A1 RNAi重组小球藻口服喂饲对白纹伊蚊的致死作用

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1. 海南医学院基础医学与生命科学学院分子生物学重点实验室, 海南海口 571199;
2. 中国热带农业科学院热带生物技术研究所/海南热带农业资源研究院, 海南海口 570216

黄小丹,女,硕士,从事热带病防治研究,E-mail:1824574713@qq.com

收稿日期: 2022-10-23

网络出版日期: 2023-06-16

基金资助

国家自然科学基金（31870344，82260669）；海南省重点研发项目（ZDYF2022SHFZ314）；海南省研究生创新科研课题（Hys2020-364）

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Lethal effects of recombinant CYP307A1 RNAi Chlorella by feeding on Aedes albopictus

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1. Key Laboratory of Molecular Biology, School of Basic Medicine and Life Sciences, Hainan Medical University, Haikou, Hainan 571199, China;
2. Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Science/Hainan Tropical Agricultural Resources Research Institute, Haikou, Hainan 570216, China

Received date: 2022-10-23

Online published: 2023-06-16

Supported by

National Natural Science Foundation of China (No. 31870344, 82260669); Key R&D Projects in Hainan Province (No. ZDYF2022SHFZ314); Hainan Postgraduate Innovative Scientific Research Project (No. Hys2020-364)

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摘要

目的了解沉默白纹伊蚊蜕皮激素合成途径酶基因CYP307A1对伊蚊幼蚊生长发育的影响,为蚊媒传染病的防控提供新的思路。方法利用RNA干扰（RNAi）技术,构建CYP307A1的RNAi表达载体,以小球藻为受体进行电击转化,筛选得到的重组藻株喂食白纹伊蚊幼蚊,使用WPS 2020软件统计并汇总幼蚊的死亡率、化蛹率、羽化率、体长和CYP307A1基因表达量实时荧光定量PCR数据；SPSS 25软件进行各组平均值的单因素ANOVA检验,使用Origin 2019和WPS 2020软件进行图表的制作。结果 300只规模实验结果显示,对照组中饲喂过滤水、饲料、野生小球藻和转pMaa7/IR载体小球藻的白纹伊蚊幼蚊死亡率分别为1.50%、0.17%、0.83%和8.00%,而饲喂CYP307A1 RNAi重组小球藻幼蚊死亡率为80.33%,死亡率显著高于各对照组,且差异有统计学意义（P<0.05）。饲喂饲料的伊蚊幼蚊化蛹率为99.83%,羽化率为98.50%,而饲喂CYP307A1 RNAi重组小球藻幼蚊化蛹率为18.50%,羽化率为98.50%,与对照组相比饲喂CYP307A1 RNAi重组小球藻幼蚊的化蛹率和羽化率显著下降,差异有统计学意义（P<0.05）。喂食重组藻株伊蚊幼蚊体内CYP307A1基因表达量比对照组下降79.00%（P<0.01）。结论 CYP307A1 RNAi重组小球藻具有抑蚊功能,有望在生物杀蚊领域发挥作用。

关键词： 白纹伊蚊; CYP307A1; RNA干扰; 小球藻

本文引用格式

黄小丹, 肖洒, 贺长皓, 李智杰, 张秀霞, 李亚军, 费小雯, 邓晓东 . CYP307A1 RNAi重组小球藻口服喂饲对白纹伊蚊的致死作用[J]. 中国媒介生物学及控制杂志, 2023 , 34(3) : 336 -343 . DOI: 10.11853/j.issn.1003.8280.2023.03.010

Abstract

Objective To investigate the effects of silencing the ecdysone synthesis pathway enzyme gene CYP307A1 of Aedes albopictus on the growth and development of Aedes larvae, and to provide new ideas for the prevention and control of mosquito-borne infectious diseases.Methods An RNAi expression vector targeting CYP307A1 was constructed using RNAi technology. Chlorella was used as the recipient for electrotransformation. The selected recombinant Chlorella strains were fed to Ae. albopictus larvae. WPS 2020 software was used to pool data on the mortality, pupation rate, emergence rate, body length, and quantitative real-time PCR results of the expression of CYP307A1 gene in larvae. One-way analysis of variance was performed on the mean values of all groups using SPSS 25. Origin 2019 and WPS 2020 were used to make charts.Results The 300-mosquito experiment showed that the mortality rates of larvae in the control groups fed with filtered water, feed, wild Chlorella, and Chlorella transferred with pMaa7/IR vector were 1.50%, 0.17%, 0.83%, and 8.00%, respectively; the mortality rate of larvae fed with recombinant CYP307A1 RNAi Chlorella was 80.33%, which was statistically higher than that of the control groups (P<0.05). The pupation rate and emergence rate of Aedes larvae fed with feed were 99.83% and 98.50%, respectively, while those of the group fed with recombinant CYP307A1 RNAi Chlorella were 18.50% and 98.50%, respectively. The pupation rate and emergence rate with recombinant CYP307A1 RNAi Chlorella were statistically lower than those of the control group (P<0.05). The expression of the CYP307A1 gene in Aedes larvae fed with the recombinant Chlorella was decreased by 79.00% compared with the control group (P<0.01).Conclusion Recombinant CYP307A1 RNAi Chlorella has anti-mosquito function, which shows promise in the field of biological mosquito control.

Key words： Aedes albopictus; CYP307A1; RNA interference; Chlorella

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