胭脂花及小报春钙调素类似蛋白基因CML19的克隆与表达分析
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引用本文:司未佳,卢婉佩,刘 颖,王 佳,程堂仁,潘会堂.胭脂花及小报春钙调素类似蛋白基因CML19的克隆与表达分析[J].西北植物学报,2019,39(4):603~612
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司未佳,卢婉佩,刘 颖,王 佳,程堂仁,潘会堂* (花卉种质创新与分子育种北京市重点实验室国家花卉工程技术研究中心城乡生态环境北京实验室北京林业大学 园林学院北京 100083) 
基金项目:中央高校基本科研业务费专项资金(2015ZCQ YL 03);
中文摘要:CML是一类重要的Ca2+响应蛋白,为了研究CML蛋白对报春花自交不亲和性的影响,该研究从胭脂花和小报春转录组数据中分别筛选出PmCML19和PfCML19基因,并对这2个基因进行全长克隆、生物信息学分析以及基因亚细胞定位和表达模式分析。结果发现:(1) PmCML19和PfCML19基因全长均为435 bp,所编码蛋白的氨基酸相似度达80.56%,均为酸性亲水性蛋白,包含4个EF手性结构;同源分析表明, PmCML19和PfCML19的亲缘关系与中华猕猴桃AcCML19最近,而拟南芥家族中的AtCML40与这2个蛋白的氨基酸序列相似度分别为44%和46%。(2)亚细胞定位结果表明,PmCML19和PfCML19基因在细胞核和细胞膜上均表达。(3)实时荧光定量PCR结果表明,PmCML19和PfCML19基因在花药中的表达量均高于雌蕊;在胭脂花和小报春亲和授粉组合的雌蕊中具有较高的表达量,而在不亲和授粉组合的雌蕊中表达量显著降低。研究推测,胭脂花和小报春CML19基因与报春花的自交不亲和性密切相关,为进一步研究异型自交不亲和遗传机制提供了新思路。
中文关键词:报春花属  钙调素类似蛋白  自交不亲和  生物信息学分析
 
Cloning and Expression Analysis of Calmodulin Like Protein Gene CML19 in Primula maximowiczii and P. forbesii
Abstract:CML is an important Ca2+ response protein. In order to investigate the effect of CML protein on self incompatibility in Primula, we screened PmCML19 and PfCML19 genes from Primula maximowiczii and P. forbesii respectively. Full length cloning, bioinformatics analysis, subcellular localization and expression pattern analysis of the two genes were performed. The results showed as follow: (1) the full length of PmCML19 and PfCML19 genes were 435 bp, and the similarity of their amino acid was 80.56%. Both of them were acid hydrophilic proteins and had four EF hand motifs. Homology analysis showed that the closest relationship with PmCML19 and PfCML19 was AcCML19 of Actinidia chinensis, while the similarity of amino acid sequences between PmCML19, PfCML19 and AtCML40 were 44% and 46% respectively. (2) The results of subcellular localization indicated that both PmCML19 and PfCML19 genes were expressed in the nucleus and cell membrane. (3) Real time fluorescence quantitative PCR results demonstrated that the expression of PmCML19 and PfCML19 in anther were much higher than that in pistil. And the expression of PmCML19 and PfCML19 in compatible pollination pistils was higher than that in incompatible pistils. Based on these results, we speculated that CML19 gene of P. maximowiczii and P. forbesii is closely correlated with self incompatibility in Primula, which provides a new idea for studying the genetic mechanism of heterotypic self incompatibility.
keywords:Primula  calmodulin like protein  self incompatibility  bioinformatics analysis
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