孤基因存在于所有已经完成测序的基因组中，具有无法在其他物种或谱系中找到序列相似基因的特点。项目前期通过芯片分析获得了三个大白菜雄性不育系可育植株花蕾特异性表达的孤基因（Bra039133, Bra023194和Bra015539）。进一步的共表达和GO富集分析显示其可能参与了花粉的发育过程。为明确其功能及探讨孤基因在花粉发育中的作用。本项目将运用qRT-PCR、启动子融合表达等技术明确三个孤基因的切确表达位置，通过构建过表达、RNAi抑制及Crispr/Cas9敲出载体进行遗传转化，分析转化植株的形态学、孢粉学、生理特性及代谢产物等的差异，明确此三基因在花粉发育中的功能。研究结果将为后续大白菜中的孤基因功能研究提供参考，丰富对孤基因的认知，丰富对花粉发育过程分子机理的理解，为人为控制大白菜育性，人为创建雄性不育体系提供新的基因源。

All sequenced genomes contain orphan genes, which exhibit no sequence similarity to any genes in other species or lineages. Previously, three orphan genes, Bra039133, Bra023194 and Bra015539, have been isolated from Br300K microarray analysis in Chinese cabbage (Brassica rapa ssp. pekinensis), which showing specifically expressed in fertile plant floral buds. Furthermore, co-expression analysis indicated all of them involved in pollen development process. To characterize the functions of these three genes during pollen development in Chinese cabbage, the qRT-PCR, promoter fusion and in stiu hybridization will be used to determine the precise expression pattern of targeted genes. Then over-expression, RNAi and Crispr/Cas9 technologies will be used for charactering the functions of targeted gene during pollen development. Morphological, palynological, physiological and metabonomical difference of the transgenic plants will be examined to analyze the biological functions of targeted genes. This research will provide a reference for the functional analysis of orphan genes in Chinese cabbage, increase our knowledge of orphan genes and the understanding of molecular mechanisms of pollen development, and provide the new gene source for artificial control of the male fertility for Chinese cabbage and creating new male sterility system for breeding.