遗传 ›› 2006, Vol. 28 ›› Issue (11): 1428-1434.doi: 10.1360/yc-006-1428

• 研究报告 • 上一篇    下一篇

白菜雄性不育相关基因BcMF4基因功能的RNAi验证

刘乐承1,2; 向 珣1; 曹家树1   

  1. 1.浙江大学蔬菜研究所, 杭州310029; 2.长江大学园艺园林学院, 荆州 434025

  • 收稿日期:2006-02-14 修回日期:2006-04-24 出版日期:2006-11-05 发布日期:2006-11-05
  • 通讯作者: 曹家树

BcMF4 Gene, Encoding a Lucine-Rich Repeat Protein, Plays a Role in Male Fertility in Chinese Cabbage-pak-choi

LIU Le-Cheng1,2; XIANG Xun1; CAO Jia-Shu1

  

  1. 1. Institute of Vegetable Science, Zhejiang University, Hangzhou 310029, China; 2. College of Horticulture and Gardening,
    Yangtze University, Jingzhou 434025, China
  • Received:2006-02-14 Revised:2006-04-24 Online:2006-11-05 Published:2006-11-05
  • Contact: CAO Jia-Shu

摘要:

BcMF4(Brassica campestris Male Fertility 4)是前一阶段从普通白菜 (Brassica campestris ssp. chinensis var. communis, syn. B. rapa ssp. chinensis var. communis)核雄性不育两用系的可育株中分离到的雄性不育相关基因。本研究根据BcMF4基因的cDNA序列设计两对特异引物, 从普通白菜花蕾cDNA中扩增出两个片断后连接至双元载体pBI12l中, 得到了RNAi (RNA interference) 植物表达栽体pBI-B4R, 并导入了农杆菌LBA4404菌株中; 通过组织培养途径转化菜心(B. campestris ssp. chinensis var. parachinensis), 72.2%的菜心转基因植株中45.8%的花粉为缩小而空瘪的畸形, 而且这些植株的花粉离体萌发率降低至23.7%; Northern杂交显示, 转基因植株的花粉畸形, 是由于BcMF4基因片段的插入使BcMF4基因的表达受到了抑制。结果表明, 采用RNAi技术下调了BcMF4基因的表达, 导致了菜心转基因植株部分花粉的不育, 证明BcMF4基因在普通白菜和菜心等白菜植物的花粉发育中起着重要作用。

关键词: 菜心, 花粉, RNAi(RNA interference), BcMF4, 普通白菜

Abstract:

The BcMF4 (Brassica campestris Male Fertility 4) gene was previously isolated from the fertile B line of Chinese cabbage-pak-choi (Brassica campestris ssp. chinensis var. communis, syn. B. rapa ssp. chinensis var. communis). In the present paper, based on the cDNA sequence of BcMF4, primers were designed and used to amplify two fragments from the cDNA of flower buds of Chinese cabbage-pak-choi. Two produced fragments were introduced separately into binary vector pBI121 in antisense and sense orientations. The generated RNA interference (RNAi) vector was then mobilized into Agrobacterium tumefaciens strain LBA4404. The A. tumefaciens harboring the BcMF4 fragments was transformed to flowering Chinese cabbage (B. campestris ssp. chinensis var. parachinensis) via tissue culture. Approximately 45.8% of the pollen grains from 72.2% of RNAi plants exhibited abnormal in their shapes, and only 23.7% of the pollen grains from these plants germinated normally. Northern blotting demonstrated that the phenotypic change of pollen grains resulted from the inhibition of expression of the BcMF4 due to the insertion of the transgene. This indicates that functional interrupting of BcMF4 by RNAi resulted in partial pollen abortion in flowering Chinese cabbage, suggesting that the product of BcMF4 gene plays an important role during pollen development of Chinese cabbage such as Chinese cabbage-pak-choi and flowering Chinese cabbage.

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