海岛棉24号染色体上粉状茸毛起始基因的精细定位及其候选基因鉴定

The initiation pattern from epidermal cells of both cotton stem trichome and cotton fibers are similar and confirmed to be regulated by analogous signal-controlled metabolic pathways. Therefore, illustration of the trichome genetic foundation will play a critical role in revealing the molecular mechanism of the cotton fiber generation and development. Cotton stem trichome can be classified into four types after systematic phenotype observation. Several F2 segregating populations with different trichome types were created in our laboratory. Using these populations, a major gene related to powdery trichome was mapped on the Chr.24(D) of Gossypium barbadense. Based on these results, the major gene of powdery trichome will be mapped finely by development of new molecular markers using published whole genome sequence of G. raimondii in this proposal,. The candidate genes will be predicted by gene annotation, and cloned from cotton varieties/accessions with different types of trichome. Association analysis will be conducted between DNA sequence features and trichome phenotype, especially with and without trichome. Candidate gene spatiotemporal expression will be quantified by real-time qPCR, which will be used to indicate their relationship with the trichome starting time and phenotype. In addition, these genes will be transformed into Arabidopsis thaliana to further verify their function.

研究表明棉花茎秆表皮茸毛的产生与棉纤维的生长发育具有相似的植物学特性，受到类似的信号代谢途径调控。因此，明确棉花茎秆茸毛产生的遗传规律对于揭示棉花纤维生长发育的分子机理具有一定的指导作用。本实验室在前期对海岛棉和陆地棉茸毛表型系统观察和科学分类的基础上，创建了不同茸毛类型的F2分离群体，并利用这些群体在海岛棉24号染色体上定位到一个粉状茸毛起始相关的主效基因。本项目基于这一初定位结果，以已发表的全基因组序列为参考，开发新的分子标记，加密遗传图谱，利用已创建的作图群体对海岛棉24号染色体上粉状茸毛基因进行精细定位。通过基因注释预测可能的候选基因并从不同茸毛类型的棉花材料中克隆，分析其DNA序列差异与茸毛表型（有无）的关系（关联分析），并根据该基因在不同时空中的定量表达推测它与茸毛起始时间及类型的相关性。此外，候选基因通过拟南芥转化实验，进一步验证其功能。

植物茸毛是表皮毛分化发育的一个分支，与纤维和根毛的发育存在相似性。本项目为寻找棉花茎秆茸毛发育的调控基因做了不懈的努力，为完善植物表皮毛发育调控机制贡献一份力量。本研究通过对多个遗传群体的调查，进一步明确24号染色体端部存在一个控制茸毛密度和起始的QTL/gene。通过对不同茎秆茸毛类型材料的基因组测序、茎秆转录组测序并结合NCBI数据库公布的棉花转录组数据等预测出基因Gh_D08G0108为最可能的候选基因。茎秆转录组数据显示目标区段内仅有Gh_D08G0108的表达量在有/无茸毛陆地棉组间的差异极显著。基于单链构象多态性（SSCP）技术，我们在130kb的目标区段内共开发了23个具有多态性的分子标记，通过一套半野生陆地棉和现代陆地棉种内杂交的F2分离群体及其F2:3群体在目标区段精细定位到了调控陆地棉茎秆茸毛起始的两个候选基因。结合生信数据分析预测，最终明确候选基因为Gh_D08G0108。Gh_D08G0108存在可变剪接，转录非编码长链RNA（lncRNA）在无茸毛品系的茎秆和叶片中特异性高表达。Gh_D08G0108基因与拟南芥同源基因GIR1或GIR2共具有高保守域，GIR1或GIR2负向调控表皮毛发育，Gh_D08G0108的lncRNA表达模式与他们一致。此外，还发现Gh_D08G0108基因第4个碱基(G/T)与茎秆无茸毛的表型高度关联。这是首次发现lncRNA参与调控棉花茎秆茸毛起始，为植物表皮毛发育调控机制奠定了新的理论基础。