Apimedica: Genetic Basis for High Royal Jelly Production

Apimondia Apimedica-Apiquality International Forum

Oct. 22-25, 2012

Zhenjiang, China

Screening on Genetic Markers Relating to Trait of High Royal Jelly Production Using Gene Chip

Jiao Pan1, Wenfeng Li, Zhiguo Li, Limin Yan, Shenglu Chen, Songkun Su,*

College of Animal Sciences, Zhejiang University, Hangzhou, China

College of Bee Science, Fujian Agriculture and Forestry University, Fuzhou, China
Contact: susongkun@zju.edu.cn

Royal jelly, a principal food of the honeybee queen and larva under 3 days old, is secreted by the hypopharyngeal and mandibular glands of nursing bees. Royal jelly has peculiar effects on health care owning to its highly active nutrients. Through years of directional selection and group closed propagation breeding, we have successfully cultivated a bee breed of high royal jelly yield, which enable China's both royal jelly production and export volume rank first in the world. So, the researches on genes related to high royal jelly producing traits and their molecular mechanisms have important theoretical significance and promising prospects in practice.

In this study, we cultivated four colonies of high royal jelly yielding bees(HRJB) and low royal jelly yielding bees (LRJB) respectively (F1,16= 66.04, P = 0.0002) by directional selection breeding. A new gene chip Agilent of honeybee covering 11689 genes (9792 genes retrieved from NCBI, 1722 newly tested genes and 175 genes which might be closely related to the royal jelly secretion) was designed and prepared, and hybridized with cDNA of nursing bees collected when they were feeding the larva in queen cells. In total, 369 differentially expressed genes between HRJB and LRJB were detected, and 201 genes were up-regulated (1.70%) in HRJB, while 168 genes were down-regulated (1.43%) in HRJB.

The cluster analysis, gene ontology (GO) and pathway of 369 differently expressed genes between HRJB and LRJB were analyzed using bio-informatics software and database. GO analyses showed that the differently expressed genes mainly participated in four biological processes including developmental regulation of organic tissue, synthesis and deposit of nutrients, oxidoreduction coenzyme and glucose metabolism, biosynthesis and metabolism of organic acid. They mainly have the following molecular functions: neuropeptide and its' receptor activity, metabolism enzymes activity of various nutrients. Pathway analyses revealed that the differently expressed genes mainly belong to 46 pathways, which can be classified into signal pathway, synthesis and metabolism of nutrients pathway, processing and modification of protein pathway.

Furthermore, 10 different colonies of bee samples were collected from royal jelly producing colonies in field, 5 of which belonged to HRJB, and another 5 were LRJB. Ten of the differentially expressed genes obtained above were further confirmed using qRT-PCR and correlation analysis, 3 genes were successfully verified to be the molecular markers relating to high royal jelly production trait. This study provided genetic basis for further studies on molecular mechanisms of high royal jelly production.

Acknowledgments

The research was supported by earmarked funds for Modern Agro-industry Technology Research System (No. CARS-45-KXJ3) and Nature and Science Foundation Commission of Zhejiang Province (R3080306) to SKS.

采用基因芯片技术筛选王浆高产性状相关的

分子标记

潘娇¹，李文峰¹，李志国¹，晏励民¹，陈盛禄¹，苏松坤^1,2*

1. 浙江大学动物科学学院  杭州  中国

2. 福建农林大学蜂学学院  福州  中国

摘要：蜂王浆（royal jelly）是由适龄工蜂发达的咽下腺和上颚腺分泌的，用于饲喂蜂王及3日龄以内幼虫，含有丰富的功能组分，对人类医疗、保健等具有独特功效。通过多年的定向选育与集团繁育，我国成功培育出王浆产量世界领先的优良王浆高产蜂种。以王浆高产蜂种为素材，研究王浆高产性状相关的基因水平的分子标记及王浆高产分子机制具有重要的理论意义和应用价值。

本研究通过定向选育手段，在同一蜂种中分别培育了王浆高产和王浆低产蜜蜂(x vs y, F_1,16= 66.04, P = 0.0002)。根据从NCBI上检索的9792条已注释的蜜蜂基因、本实验室在蜜蜂幼虫中新检测的1722条基因和175条与王浆分泌可能紧密相关的基因，设计和制备了蜜蜂基因芯片（Agilent），分别与王浆高产蜜蜂（HRJB）及王浆低产蜜蜂(LRJB)中正在吐浆的哺育蜂的头部cDNA进行杂交，初步筛选出369条差异表达的基因，其中在王浆高产蜜蜂中上调表达的基因有201个（1.70%），下调表达168个（1.43%）。

运用高级生物信息学分析软件对基因芯片所得到的369个差异表达基因进行聚类分析、GO分析和Pathway分析。结果表明，差异表达基因主要参与了器官组织的发育调控过程、营养物质的合成与沉积过程、氧化还原辅酶与葡萄糖的代谢过程和有机酸的合成与代谢过程；分子功能主要为神经肽及受体活性、物质代谢酶活性；参与信号通路、营养物质合成代谢、蛋白质加工及修饰代谢途径共有46个。

此外，采集王浆高、低产蜜蜂样本，通过qRT-PCR技术和相关性分析对其中的10个差异表达基因进行验证，成功筛选出3个基因可以作为王浆高产性状相关的分子标记。这些研究结果为揭示王浆高产的分子机制奠定了良好基础。

关键词：蜜蜂(Apis mellifera)；王浆产量性状；基因芯片；qRT-PCR 分子标记