兰州熊蜂（Hymenoptera:Apidae）雌性蜂产卵调控的分子机制

Bombus lantschouensis is a native and excellent bumblebee species in China because its pollination performance and potential for factory reproduction. The female reproductive status and regulation of bumblebee will determine the fate of bumblebee colony, which is the essential factor for artificial utilization of bumblebee. Therefore, to explore the molecular regulation mechanisms of female bumblebee has important practical significance. Previous studies have shown that the status of ovarian development will directly affect laying eggs by female bumblebee, especially the development of worker ovary will affect the lifespan of colony. The differential gene expression in ovary between egg-laying and non-egg laying queens and workers will be explored by using the transcriptome sequencing. The full length of key genes will clone with rapid-amplification of cDNA ends (RACE) method. The expression pattern of egg laying related genes will measure by using RT-qPCR and western blotting. Finally, the function of key genes will be validated by RNAi technology. Our project will use high-throughput sequencing technology to explore molecular mechanism of oviposition regulation in female bumblebee. The reproductive biology of bumblebee will be promoted from this project. Moreover, this project will help to improve the technology of artificial bumblebee rearing level.

兰州熊蜂（Bombus lantschouensis）以其群势大、易饲养和授粉性能优良等优点成为我国人工繁育重点攻关蜂种。雌性蜂的生殖调控直接影响蜂群建立和发展，是熊蜂繁育的重要影响因素，因此，探明雌性蜂产卵调控的机制具有重要的现实意义。前期研究表明，卵巢发育程度直接影响雌性蜂产卵，特别是工蜂卵巢的发育会影响蜂群寿命。本项目拟以兰州熊蜂雌性蜂卵巢为材料，利用高通量测序方法对蜂王和工蜂产卵前后卵巢组织差异的转录表达进行分析，以期探明雌性蜂产卵前后卵巢组织的基因表达差异；同时，利用RACE方法克隆主要调控基因，并利用荧光定量PCR与蛋白免疫印迹的方法分析关键基因表达特性；最后，采用RNAi技术验证调控产卵的关键基因功能。本项目利用高通量测序技术探索熊蜂雌性蜂产卵调控的分子机制，以期为熊蜂生殖生理研究奠定理论基础，也为熊蜂工厂化繁育提供指导。

本项目重点对兰州熊蜂雌性蜂-工蜂产卵调控的分子机制进行了研究，并筛选了雌性蜂产卵相关的主要基因，获得主要结果如下：（1）通过对产卵与未产卵工蜂血淋巴蛋白质组和卵巢转录组分析研究，发现工蜂产卵后机体抗氧化、免疫和解毒能力增强，能量代谢和蛋白合成维持较高水平，Notch信号通路和Hippo信号通路等信号转导途径可能在熊蜂工蜂产卵调控中起着重要的作用，并寻找到产卵调控相关的差异表达基因；（2）筛选获得兰州熊蜂雌性蜂产卵调控主要基因IRP30及其表达情况，明确了IRP30的基因结构及系统进化情况，同时发现IRP30蛋白在熊蜂发育和产卵过程中均高表达，表明该蛋白不仅能响应免疫，还可能在变态发育和繁殖过程中发挥重要作用，具有促进蜂王产卵和抑制工蜂产卵的应用前景。本项目的完成为实现人工抑制工蜂产卵现象以延长熊蜂蜂群使用寿命提供理论基础，对提高我国本土熊蜂人工周年繁育技术，促进我国熊蜂授粉产业的发展具有重要意义。