不结球白菜调控开花时间候选基因BcVIL1的克隆与表达分析

[目的]本文旨在克隆和研究不结球白菜BcVIL1基因在春化过程中的表达情况，初步了解不结球白菜BcVIL1基因的结构及功能。[方法]以普通白菜PC-175和菜心‘CX-49’为试验材料，利用RT-PCR技术克隆BcVIL1基因全长，并采用生物信息学方法进行氨基酸序列比对和进化分析。通过实时荧光定量PCR（RT-qPCR）技术分析2种材料春化过程中BcVIL1的表达情况。构建亚细胞定位载体pEZS-NL-BcVIL1，利用亚细胞定位技术，分析BcVIL1基因在真核细胞中表达的位置。[结果]2种材料中BcVIL1基因序列基本一致，开放阅读框为1 020 bp，编码339个氨基酸。进化分析表明：BcVIL1基因与大白菜进化关系最近，同源性最高（99%）。在PC-175中，春化处理7 d就可以诱导BcVIL1基因的表达，且在整个春化过程及春化之后都有较高的水平表达，而在‘CX-49’中，BcVIL1基因在春化过程中的表达与未春化处理相比无显著差异。亚细胞定位结果表明：BcVIL1主要在细胞核与细胞膜上表达。[结论]BcVIL1基因在春化过程中其表达具有品种特异性，且在不结球白菜中该基因主要在细胞核与细胞膜上发挥作用。
[Objectives] BcVIL1 gene had been cloned and some experiments had been producted to better understand the characteristics and structure of BcVIL1.[Methods]To explore the function of the VIL1 gene in non-heading Chinese cabbage,PC-175 and ‘CX-49’ were chosen as materials which belong to typical different genotypes. The BcVIL1 gene in PC-175 and ‘CX-49’ was cloned by RT-PCR,and bioinformatics analysis was conducted. The expression patterns of BcVIL1 in PC-175 and ‘CX-49’ during the course of vernalization were analyzed by RT-qPCR. The vector pEZS-NL-BcVIL1 was constructed and the spatial expression of BcVIL1 was analyzed by sub-cellular localization.[Results]The sequences of BcVIL1 were consistent in two materials of non-heading Chinese cabbage,and BcVIL1 containing a 1 020 bp open reading frame encoded 339 amino acid residues. The bioinformatics analysis showed that the homology between Brassica campestris ssp. chinensis Makino and B. rapa ssp. pekinensis was very high at 99%. The BcVIL1 gene in PC-175 could be induced by 7 days of vernalizaing and the expression of BcVIL1 gene was at high level during and after the course of vernalization. While the expression of BcVIL1 gene in ‘CX-49’ had no obvious differentce compared with non-vernalization condition. The BcVIL1 gene was verified to be located in cell nucleus and membrane by sub-cellular localization.[Conclusions]The vernalization in PC-175 can induce the expression of BcVIL1,and the expression of BcVIL1 also was at high level after vernalization. However the expression of BcVIL1 gene in ‘CX-49’ didn’t have this profile. The expression of BcVIL1 gene was different in two genotypes. BcVIL1 gene in non-heading Chinese cabbage mainly had a function at cell nucleus and membrane