小报春类钙调素蛋白基因PfCML41的克隆及序列分析

Cloning and Sequence Analysis of Calmodulin-like Gene PfCML41 from Primula forbesii

CMLs是一类钙离子传感器,能与Ca2+结合,对促进花粉萌发和花粉管生长发挥重要作用。为了探究CMLs对小报春(Primula forbesii)异型自交不亲和系统的影响,在小报春以长花柱做母本时的亲和授粉与不亲和授粉的转录组数据中发现了一个差异表达基因PfCML41,并对其进行了CDS序列全长克隆和基因序列分析。结果表明,PfCML41基因的CDS序列全长为555 bp,编码184个氨基酸。编码的蛋白无信号肽和跨膜区域,属于亲水性蛋白。PfCML41含有EF-hand结构,能与Ca^(2+)结合。通过系统进化树分析,PfCML41与茶(Camellia sinensis)的CsCML41亲缘性更近。实时荧光定量PCR结果表明PfCML41在不亲和授粉组合中的相对表达量高于亲和授粉组合,且差异显著。本研究为小报春PfCML41的表达分析与功能验证提供了理论依据。

CMLs, a kind of calciumion sensors, can combine with Ca2+, which plays an important role in the process of pollen germination and pollen tubes growth. In order to explore the effects of CMLs on heteromorphic self-incompatibility system of Primula forbesii, a new differential expression gene PfCML41 was found through filtering in the database of transcriptome sequencing, which compared compatible and incompatible pollination on P. forbesii when long style was used as female parent. The full-length cloning(CDS) of PfCML41 and its sequence analysis was carried out. The results showed that the full-length CDS sequence of PfCML41 was 555 bp, encoding184 amino acids. The protein had no signal peptide and transmembrane region, and belonged to hydrophilic protein. PfCML41 contained EF-hands, which can be combined with Ca^(2+). By phylogenetic tree analysis, PfCML41 had a closer homology with CsCML41 of Camellia sinensis. Quantitative real-time PCR results showed that the relative expression of PfCML41 in incompatible pollination combinations was higher than that in compatible pollination combinations, and the difference was significant. This study provided a theoretical basis for the expression analysis and functional verification of P. forbesii gene PfCML41.