DNA甲基化在蜂王发育过程中的调控作用

The queen is the core of a honeybee colony. Its quality strongly relates to the dynamics and productivity of the whole colony, and may also potentially relates to the colony collapse disorder.. DNA methylation is one of epigenetic phenomena. To explore effects of DNA methylation on the regulation of honeybee queen development, this study plan to use fertilized eggs, 1.5d and 3d worker larvae to rear queens continuously for 5 generations. Afterwards, queen morphological indexes such as weight and ovariole number will be tested, and the gene express and DNA methylation of five queen-quality related genes dynactin p62,Vg,JH,defensin-1 and CytP450 will be compared among each treatment and generation. This allows to explore the relationship among queen quality, DNA methylation and gene expression, and also to explore whether the altered DNA methylation sites could input into next generation and it effects could be accumulated by generations. This could enormously contribute to our understanding on the mechanism of how DNA methylation regulates on honeybee queen development. Carrying out this project will offer a theoretical support for high-quality honeybee queen rearing, and is expected to provide new ideas and methods to protect and utilize the valuable resource of this main pollinator, which has important theoretical and practical significance.

蜂王是蜂群的核心，蜂王的质量优劣不仅直接关系到蜂群群势强弱以及蜂产品产量高低，而且可能与“蜂群衰竭失调病”相关。. DNA甲基化是一种常见的表观遗传学现象。为了阐明DNA甲基化在蜂王发育过程中的调控作用，本研究以西方蜜蜂为实验材料，以卵、1.5日龄幼虫、3日龄幼虫三种方法进行连续多代培育蜂王，测定蜂王形态质量指标（包括初生重，卵巢管数等），同时选定与蜂王质量发育相关以及甲基化有差异的dynactin p62、Vg、JH、defensin-1、CytP450五个基因进行甲基化位点分析，并对这五个基因进行定量表达检测，以寻找蜂王质量、基因甲基化、基因定量表达三者之间内在联系，同时探索蜜蜂甲基化能否母代遗传以及累代效应，进而揭示DNA甲基化在蜂王发育过程中的调控机制。项目研究结果不仅为优质蜂王培育可提供理论支撑，而且对蜜蜂保护和利用有望提供新思路与方法，因此具有重要理论和实际意义。

为了阐明DNA甲基化在蜂王发育过程中的调控作用，本研究以西方蜜蜂为实验材料，移工蜂巢房中卵、1日龄幼虫、2日龄幼虫和3日龄幼虫进行人工培育蜂王，测定与蜂王发育相关指标。对5个蜂王发育相关基因14个DNA片段进行单基因甲基化检测。结果表明：随着移虫日龄增加，培育蜂王的质量下降；但4个实验组5个基因的14个DNA片段的甲基化水平都差异不显著。. 利用Illumina高通量测序平台对蜂王样品进行转录组测序，结果表明：随着移虫日龄的增加，移虫培育的蜂王与移卵培育的蜂王之间的差异表达基因数量逐步增多。利用全基因组Bisulfite甲基化测序技术，对人工培育的蜂王进行全基因组DNA甲基化测序，结果表明：蜂王质量随着移虫日龄的增加而降低；随着移虫日龄的增加，移虫培育的蜂王与移卵培育的蜂王之间的差异甲基化基因数量逐步增多。利用用全基因组Bisulfite甲基化测序技术，对4代人工培育的蜂王进行全基因组DNA甲基化测序。结果表明：四代蜂王中均是E组育王方式培育的蜂王质量最好，L2组最差；还发现：蜜蜂DNA甲基化具有累代效应。利用全基因组 Bisulfite甲基化测序技术，对G0代蜂王、G1代蜂王、G2代雄蜂进行全基因组DNA甲基化测序，结果表明：DNA甲基化遗传印迹能够在蜜蜂这一模式生物体内稳定遗传。研究结果不仅为优质蜂王培育可提供理论支撑，而且对蜜蜂保护和利用有望提供新思路与方法，因此具有重要理论和实际意义。