Journal of Integrative Agriculture ›› 2014, Vol. 13 ›› Issue (5): 954-962.DOI: 10.1016/S2095-3119(13)60581-X

• 论文 • 上一篇    下一篇

Decreased Pollen Viability and Thicken Pollen Intine in Antisense Silenced Brassica campestris Mutant of BcMF19

 LIU Jin-long, GAO Ming-hui, LIU Ying , CAO Jia-shu   

  1. Laboratory of Cell and Molecular Biology, Institute of Vegetable Science, Zhejiang University, Hangzhou 310058, P.R.China
  • 收稿日期:2013-03-20 出版日期:2014-05-01 发布日期:2014-05-06
  • 通讯作者: CAO Jia-shu, Tel/Fax: +86-571-88982188, E-mail: jshcao@zju.edu.cn
  • 基金资助:

    This work was supported by the National Basic Research Program of China (2012CB113900), the National Natural Science Foundation of China (31071805), and the Key Sci-Technology Project of Zhejiang Province, China (2010C12004).

Decreased Pollen Viability and Thicken Pollen Intine in Antisense Silenced Brassica campestris Mutant of BcMF19

 LIU Jin-long, GAO Ming-hui, LIU Ying , CAO Jia-shu   

  1. Laboratory of Cell and Molecular Biology, Institute of Vegetable Science, Zhejiang University, Hangzhou 310058, P.R.China
  • Received:2013-03-20 Online:2014-05-01 Published:2014-05-06
  • Contact: CAO Jia-shu, Tel/Fax: +86-571-88982188, E-mail: jshcao@zju.edu.cn
  • Supported by:

    This work was supported by the National Basic Research Program of China (2012CB113900), the National Natural Science Foundation of China (31071805), and the Key Sci-Technology Project of Zhejiang Province, China (2010C12004).

摘要: Brassica campestris male fertility 19 (BcMF19; GenBank accession number GQ902048.1), a gene that is specially expressed in tapetum cells and microspores during anther development in B. campestris ssp. chinensis, which is learned from the previous in situ hybridization study. In the present study, we constructed antisense-silenced plants of BcMF19 using B. campestris ssp. chinensis to validate this prediction. The morphology of the pistils, long anthers, and short anthers was significantly affected in 35sbcmf19 compared with the control samples. 4´-6-Diamidino-2-phenylindole staining revealed that two generative nuclei and one large vegetative nucleus were not affected in the mutant compared with control. Statistical analysis of Alexander’s staining results showed that 96% of the control pollen grains had vitality, whereas only 86% of the mutant pollen grains did. Under scanning electron microscopy, the mutant demonstrated numerous abnormal pollen grains and resembled dried persimmon. The frequency of normal pollen grains was approximately 18%. Under transmission electron microscopy, the pollen intine during the binucleate and mature pollen stages in 35sbcmf19 exhibited abnormal thickening, especially at the germinal furrows, compared with control. In vitro pollen germination test showed that the tips of the mutant pollen tubes transformed into globular alveoli and stopped growing compared with control. On the other hand, in vivo pollen germination test suggested that BcMF19 affected the pollen tube extension in the pistil. These findings indicate that BcMF19 is essential to the pollen development and pollen tube extension of B. campestris ssp. chinensis.

关键词: Brassica rapa ssp. chinensis , PGIP , pollen , polygalacturonase-inhibiting protein , tapetum , pollen germination

Abstract: Brassica campestris male fertility 19 (BcMF19; GenBank accession number GQ902048.1), a gene that is specially expressed in tapetum cells and microspores during anther development in B. campestris ssp. chinensis, which is learned from the previous in situ hybridization study. In the present study, we constructed antisense-silenced plants of BcMF19 using B. campestris ssp. chinensis to validate this prediction. The morphology of the pistils, long anthers, and short anthers was significantly affected in 35sbcmf19 compared with the control samples. 4´-6-Diamidino-2-phenylindole staining revealed that two generative nuclei and one large vegetative nucleus were not affected in the mutant compared with control. Statistical analysis of Alexander’s staining results showed that 96% of the control pollen grains had vitality, whereas only 86% of the mutant pollen grains did. Under scanning electron microscopy, the mutant demonstrated numerous abnormal pollen grains and resembled dried persimmon. The frequency of normal pollen grains was approximately 18%. Under transmission electron microscopy, the pollen intine during the binucleate and mature pollen stages in 35sbcmf19 exhibited abnormal thickening, especially at the germinal furrows, compared with control. In vitro pollen germination test showed that the tips of the mutant pollen tubes transformed into globular alveoli and stopped growing compared with control. On the other hand, in vivo pollen germination test suggested that BcMF19 affected the pollen tube extension in the pistil. These findings indicate that BcMF19 is essential to the pollen development and pollen tube extension of B. campestris ssp. chinensis.

Key words: Brassica rapa ssp. chinensis , PGIP , pollen , polygalacturonase-inhibiting protein , tapetum , pollen germination