陆地棉Ghkinesin13亚家族基因的克隆及表达特征分析

Kinesin家族是一类马达蛋白,它们能利用ATP水解所释放的能量驱动自身携带物质分子沿着微管运动,在细胞形成、细胞伸长等方面起着关键作用。本研究以拟南芥Atkinesin-13A蛋白序列作为探针序列,利用Blast比对从二倍体雷蒙德氏棉的基因组数据库中发现7个具有较高同源关系的基因。根据基因序列设计引物,利用RT-PCR技术从陆地棉纤维中分离出7个基因。依据7个基因与Atkinesin-13A和Atkinesin-13B的同源性高低,依次将其命名为Gh KIS13A1、Gh KIS13A2、Gh KIS13A3、Gh KIS13B1、Gh KIS13B2、Gh KIS13B3和Gh KIS13B4。生物信息学分析表明,7个Ghkinesin13均含有典型的KISC马达区域、ATP结合位点和微管结合位点,其马达区域属于中央马达。多重序列比对和进化树分析发现,这7个蛋白可被分为2个(Kinesin13A和Kinesin13B)亚类。实时荧光定量PCR结果表明,7个Ghkinesin13亚家族基因在棉花各组织中均有表达,但表达模式各不相同,其中Gh KIS13B4在纤维中优势表达,表明其在纤维发育过程中可能发挥着重要作用。

Cloning and Expression Analysis of Ghkinesin13 Subfamily Genes in Gossypium hirsutum

Kinesin family belongs to a class of motor proteins. Kinesins can move along microtubule filaments by using the energy released from ATP hydrolysis, and play key roles during cell formation and cell elongation. Using the protein sequence of Atkinesin13A as a probe, seven genes with high sequence homology were obtained from a genome database of Gossypium raimondii diploid cotton with Blast alignment. These sequences were used to design primers, and then seven genes were isolated from upland cotton fiberusing RT-PCR. Based on the homology level of the seven genes with Atkinesin13A and Atkinesin13B, we designated them as GhKIS13A1, GhKIS13A2, GhKIS13A3, GhKIS13B1, GhKIS13B2, GhKIS13B3, and GhKIS13B4 respectively. Bioinformatic analysis showed that seven proteins contained typical KISC domains including the central motor, ATP-binding sites and microtubule binding sites. Multiple sequence alignment and phylogenetic tree analysis revealed that seven genes can be divided into Kinesin13A and Kinesin13B. The qPCR showed that the seven Ghkinesin13 subfamily genes expressed in all tissues of cotton, but showed different expression patterns. Of the seven genes, only GhKIS13A4 was preferentially expressed in fibers, suggesting that it may play an important role in cotton fiber development. This study can provide a foundation for studying the functions of Ghkinesin13 subfamily genes in fiber development.