水稻OsHDT703基因的可变剪接体分析

Studies on Alternative Splicing Variants of Rice OsHDT703 Gene

为了克隆水稻HD2蛋白基因OsHDT703,并分析其存在的可变剪接体。利用生物信息学分析,结合RT-PCR、TA克隆和测序等方法,鉴定1个水稻中全新的HD2蛋白成员(OsHDT703)。结构域分析发现,在其N端含有HD2家族的NPL保守结构域。选取来源于7个物种的16个HD2蛋白,构建蛋白进化树,结果发现,OsHDT703蛋白与OsHDT702蛋白处于同一分支;OsHDT701则处于另一分支,说明水稻中HD2家族蛋白可能存在功能分化。根据预测的不同剪接体设计4对特异性引物,以粳稻中花11叶片和幼穗cDNA为模板,利用RT-PCR扩增,结合TA克隆,通过测序比对,成功鉴定到4种不同可变剪接体序列。由于3号(OsHDT703.3)和4号(OsHDT703.4)剪接体的CDS序列完全相同,所以证明OsHDT703至少存在4种不同可变剪接体。综上所述,鉴定到水稻中1个全新的HD2蛋白,命名为OsHDT703,并且鉴定出至少4种存在可变剪接体,为后续深入研究该基因生物学功能奠定基础。

In the study,the alternative splicing variants of new HD2 protein(OsHDT703)were identified by bioinformatics analysis combining with RT-PCR,TA cloning and sequencing technology.A new HD2 family protein(OsHDT703)was detected by bioinformatics analysis,which contained the NPL conserved domain at N-terminus.Sixteen HD2 proteins were chosen from seven species to construct evolutionary tree.The results showed that OsHDT703 protein was in the same branch as OsHDT702 protein,and OsHDT701 was in another branch,indicating that HD2 family proteins might have the functional differentiation in rice.Four specific primers were designed based on different alternative splicing variants,and the cDNA of rice leaves was used as template.At least four alternative splicing variants of OsHDT703 were cloned by RT-PCR and sequencing.Because the CDS sequences of OsHDT703.3 and OsHDT703.4 were the same,indicating that OsHDT703 had at least 4 different alternative splicing variants.In summary,a new HD2 protein was identified in rice,named as OsHDT703,and at least four alternative splicing variants were identified,which laid the foundation for further study on the of OsHDT703 function.