辣椒胞质雄性不育系CaCOX3的克隆与表达分析

Cloning and Expression Analysis of CaCOX3 Gene from Pepper Cytoplasmic Male Sterile Line

探究线粒体呼吸链第4个复合体亚基细胞色素氧化酶第三亚基(COX3)与辣椒胞质雄性不育之间的关系,为研究辣椒CMS与能量代谢的关系提供参考。以辣椒胞质雄性不育系9704A和保持系9704B为材料,根据GenBank报道的辣椒线粒体基因组序列,设计特异引物扩增CaCOX3,进行生物信息学分析,并研究其时空表达特性,比较两系间的差异。在辣椒胞质雄性不育系9704A和保持系9704B中克隆获得的目的基因CaCOX3,全长均为798 bp,编码265个氨基酸残基;辣椒保持系9704B不同组织中,CaCOX3表达存在差异,果皮中表达最高,叶中表达最低。在不同的花蕾发育时期,两系CaCOX3表达存在差异。胞质雄性不育系9704A的造孢细胞增殖期和花粉母细胞减数分裂期,CaCOX3的表达量明显低于保持系9704B;在小孢子单核期,两系CaCOX3的表达量持平;在小孢子成熟期,胞质雄性不育系明显高于保持系。CaCOX3在辣椒胞质雄性不育9704A和保持系9704B的时空表达存在差异,可能引起能量代谢异常,造成不育。

This work aims to explore the relationship between the encoding of the third sub-unit of cytochrome oxidase in the 4th complex subunit of mitochondrial respiratory chain(COX3)and the cytoplasmic male sterility(CMS)of pepper,for providing a reference in studying the relationship between pepper CMS and energy metabolism.Pepper cytoplasmic sterile line 9704A and maintainer line 9704B were used as material,CaCOX3 gene was amplified using specific primers based on the sequence of capsicum mitochondrial genome in GenBank.Its bioinformation and spatiotemporal expression were analyzed and the difference between 2 lines was compared.The cloned target CaCOX3 gene from the 9704A and the 9704B were consistent,the whole length was 798 bp,and encoded 265 amino acid residues.The expressions of CaCOX3 gene in different tissues of the 9704B were different:the highest in pericarp and the lowest in leaves.CaCOX3 gene expressed differently during the different developing periods of the bud of both lines.CaCOX3 gene expressions in the proliferative stage of sporogenetic cells and the meiotic stage of pollen mother cells of 9704A line were significantly lower than those of 9704B.The expressions in both lines were the same at the stage of microspore monocytes.The expressions in the CMS line was significantly higher than that in the maintainer line at the microspore maturation.In sum,the spatiotemporal expressions of CaCOX3 gene were different in the CMS line 9704A and the maintainer line 9704B,resulting in the abnormal energy metabolism and then infertility.