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作物学报 ›› 2017, Vol. 43 ›› Issue (02): 201-209.doi: 10.3724/SP.J.1006.2017.00201

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

小麦TaVIP1家族基因克隆、分子特性及功能分析

赵佩1,**,腾丽杰2,**,王轲1,杜丽璞1,任贤2,佘茂云3,*,叶兴国1,*   

  1. 1中国农业科学院作物科学研究所 / 国家农作物基因资源与基因改良重大科学工程, 北京 100081; 2北方民族大学生物科学与工程学院, 宁夏银川 750002; 3安徽省农业科学院作物研究所, 安徽合肥 230031
  • 收稿日期:2016-03-25 修回日期:2016-09-18 出版日期:2017-02-12 网络出版日期:2016-09-28
  • 通讯作者: 佘茂云, E-mail: ahxiaoshe@126.com; 叶兴国, E-mail: yexingguo@caas.cn
  • 基金资助:

    本研究由国家自然科学基金项目(31401380, 31401376)资助。

Cloning, Molecular Characterization, and Functional Analysis of Wheat TaVIP1 Genes

ZHAO Pei1,**,TENG Li-Jie2,**,WANG Ke1,DU Li-Pu1,REN Xian2,SHE Mao-Yun3,*,YE Xing-Guo1,*   

  1. 1Institute of Crop Science, Chinese Academy of Agricultural Sciences / National Key Facility for Crop Gene Resources and Genetic Improvement, Beijing 100081; 2 College of Biology and Engineering, Beifang University of Nationalities, Yinchuan 750002, China; 3Crop Research Institute, Anhui Academy of Agricultural Sciences, Hefei 230031, China
  • Received:2016-03-25 Revised:2016-09-18 Published:2017-02-12 Published online:2016-09-28
  • Contact: 佘茂云, E-mail: ahxiaoshe@126.com; 叶兴国, E-mail: yexingguo@caas.cn
  • Supported by:

    This study was supported by the National Natural Science Foundation of China (31401380, 31401376).

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摘要：

植物VIP1蛋白(VirE2 interacting protein 1)与农杆菌侵染植物后T-DNA在细胞内的转运有关，影响植物转化效率，但还未见有关小麦中VIP1基因研究的报道。本研究利用in silico技术从普通小麦中克隆了TaVIP1家族基因，该家族基因全部由4个外显子构成，编码产物相似性高，与拟南芥AtVIP1蛋白质序列相似性仅为50.7%~51.4%。Southern杂交结果表明，TaVIP1基因在小麦基因组中存在3个拷贝。根据小麦3个TaVIP1基因的差异序列设计特异引物，以四倍体小麦Langdon与中国春D组染色体代换系为材料进行PCR检测，结合生物信息学分析，将小麦3个小麦TaVIP1基因分别定位在4AL、4BS和4DS染色体上。亚细胞定位分析表明，TaVIP1蛋白分布于细胞膜、细胞质和细胞核中。农杆菌转化烟草，过表达TaVIP1基因降低了烟草转化效率。小麦TaVIP1基因编码的氨基酸序列与拟南芥AtVIP1基因及烟草NtVIP1基因编码的氨基酸序列相似性很低，这可能是小麦农杆菌转化效率低的原因之一。

关键词: 小麦, TaVIP1, 烟草, 农杆菌转化

Abstract:

Plant VIP1 (VirE2 interacting protein 1) is involved in the transportation of T-DNA in plant cells after Agrobacterium infection, affecting plant transformation efficiency. However, the function of TaVIP1 is unknown. Using in silico technique, a family of TaVIP1 genes were successfully cloned from common wheat in this study. The gene family encompasses four exons and shares high similarity among its members while only 50.7%–51.4% similarity to the AtVIP1 in Arabidopsis. Southern blotting assay showed that the TaVIP1 had three allelic copies in wheat genome. The three copies of TaVIP1 were further assigned to wheat chromosomes 4AL, 4BS, and 4DS according to Blastn in the IWGSC (International Wheat Genome Sequencing Consortium) database, and experimentally confirmed by PCR using specific primers to each TaVIP1 allele based on their DNA sequences and Langdon durum disomic substitution lines. Subcellular localization analysis showed that TaVIP1 proteins were located in cell membrane, cytoplasm and nucleus. Over-expression of TaVIP1 in tobacco obviously reduced Agrobacterium-mediated transformation efficiency. The amino acid sequence encoded by TaVIP1 only had less similarity to the corresponding amino acid sequence encoded by AtVIP1 in Arabidopsis or NtVIP1 in tobacco. This might be a reason of the low transformation efficiency of wheat mediated by Agrobacterium.

Key words: Wheat, TaVIP1, Tobacco, Agrobacterium-mediated transformation

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