权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Evolutionary and immunological basis of an insect intracellular symbiosis

昆虫细胞内共生的进化和免疫学基础

基本信息

批准号：
19H03275
负责人：
松浦優
金额：
$ 10.9万
依托单位：
University of the Ryukyus
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2019
资助国家：
日本
起止时间：
2019-04-01 至 2023-03-31
项目状态：
已结题

项目摘要

本研究では、細胞内共生を派生的に獲得したヒメナガカメムシとその共生細菌を主な対象に自然免疫系の遺伝子群の機能解析や細胞培養実験系を通して細胞内共生の包括的理解を目指している。本年度は遺伝子の機能解析を中心に進めた。ヒメナガカメムシ由来cDNAあるいは長鎖合成オリゴヌクレオチドを元に、それぞれ20、30種のdsRNAを合成した。それらを150 ng/ulの濃度で若い雌成虫に注射して、母性RNAiを誘導し、卵の観察と固定、共生細菌の定量を行った。十分な核酸量確保のために10個の卵を1サンプルとして抽出した。結果、３つの遺伝子処理区において、多数の成虫致死や極端な胚発生異常がみられたため、それらの遺伝子配列を検討対象から除外した。既に機能が判明しているUbxと他２つの処理区において菌細胞形成の異常が見られた。これらの２遺伝子について、追加で個別にRNAi処理を実施して、多数の卵を観察、固定した。さらに、RNAiによる菌細胞喪失に伴う共生細菌の減少量の検出が可能か、定量的PCRを実施したところ、Ubx-RNAiにより共生細菌コピー数が2から3割減少したが、値が大きくばらついたため、RNAi後の発生段階がずれやすい卵ではなく孵化直後の個体を用いて再実験を行う必要がある。また、オリゴヌクレオチド由来のRNA合成は実験操作を簡略化コスト減に成功したがRNAi効果の早期減衰が見られたため、重要な処理区ではdsRNAの配列を長くして再実験した。当初の予定年度内および繰越年度においても感染症の影響を大きく受け、実験と観察の中断が度々生じて、ベンチワークが遂行不能となったため、細胞内共生細菌Lariskellaのゲノム解析、ナガカメムシ類のミトコンドリアゲノムの多型解析、沖縄産害虫の腸内細菌多様性およびゲノム解析、昆虫共生菌・寄生菌類のゲノム解析などリモート環境で実施可能な研究を進めた。

In this study, we obtained the results that the symbiotic bacteria derived from intracellular symbiosis were similar to those of the natural immune system, which were able to analyze the understanding of intracellular symbiosis in the cell culture system. This year, the sub-machine will be able to analyze the import of the center. The reason for this is that cDNA has been synthesized for a long time, so that you can synthesize a lot of dsRNA. If the female adult was injected with 150 ng/ul, the maternal RNAi was injected, the egg was fixed, and the symbiotic bacteria were quantified. The amount of nucleic acid is 10%. Make sure that 10 eggs are extracted from each egg. Results: the results showed that there were some diseases in the pathogens of 3-day-old adults, most of the adult embryos were born at the end of the embryo, the embryos of most adults were born at the end of the embryo, the embryos of most adults were born at the end of the embryo, the embryos were born in the embryo, the embryos were born in the embryo, the eggs were born in the embryo, the embryos were born in the embryo, the eggs were born in the embryo, the eggs were born in the embryo, and the seeds were divided into two groups. The machine has been able to determine that the cells of bacteria in the Ubx area are often found to be abnormal. Please make sure that you need to pay more attention to each other, add more information about RNAi treatment, check and fix most of the eggs. The cell loss of bacteria associated with symbiotic bacteria was accompanied by a small amount of symbiotic bacteria, quantitative PCR was used to treat symbiotic bacteria, Ubx-RNAi symbiotic bacteria were used to reduce the number of bacteria, the number of bacteria was 2, the number of bacteria was 3, and the eggs were incubated. After RNAi, the eggs were incubated and the eggs were incubated. The cause of the RNA synthesis, the operation of the operation, the success of the RNAi, the early decline of the fruit, the failure of the early stage, the dsRNA of the important area, the operation of the operation, the operation of the operation, the success of the operation, the operation of the operation, the success of the operation, the failure of the early stage, the failure of the early stage, the configuration of the important area, the queue, the queue and the queue. In the first part of the year, the number of infectious diseases in the planned year, the number of infections, the degree of infection, the number of bacteria, the number of bacteria in The analysis of insect symbiotic bacteria and parasitic bacteria may improve the research on the application of the environment.