In vivo fluorescence observation of parasporal inclusion formation in Bacillus thuringiensis

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• 作者：Hui Yang (1) (2)
  Rong Rong (1)
  FuPing Song (1)
  ChangPo Sun (1)
  Juan Wei (1)
  Jie Zhang (1)
  DaFang Huang (2)
  
• 关键词：Bacillus thuringiensis ; Cry1Ac ; GFP fusion protein ; laser confocal microscopy
• 刊名：Science China Life Sciences
• 出版年：2010
• 出版时间：September 2010
• 年：2010
• 卷：53
• 期：9
• 页码：1106-1111
• 全文大小：1047KB
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• 作者单位：Hui Yang (1) (2)
  Rong Rong (1)
  FuPing Song (1)
  ChangPo Sun (1)
  Juan Wei (1)
  Jie Zhang (1)
  DaFang Huang (2)
  
  1. State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing, 100193, China
  2. Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing, 100081, China
  

文摘

A recombinant gene expressing a Cry1Ac-GFP fusion protein with a molecular mass of approximately 160 kD was constructed to investigate the expression of cry1Ac, the localization of its gene product Cry1Ac, and its role in crystal development in Bacillus thuringiensis. The cry1Ac-gfp fusion gene under the control of the cry1Ac promoter was cloned into the plasmid pHT304, and this construct was designated pHTcry1Ac-gfp. pHTcry1Ac-gfp was transformed into the crystal-negative strain, HD-73 cry?/sup>, and the resulting strain was named HD-73?/sup>(pHTcry1Ac-gfp). The gfp gene was then inserted into the large HD-73 endogenous plasmid pHT73 and fused with the 3-terminal of the cry1Ac gene by homologous recombination, yielding HD-73Φ(cry1Ac-gfp)3534. Laser confocal microscopy and Western blot analyses showed for the first time that the Cry1Ac-GFP fusion proteins in both HD-73?/sup>(pHTcry1Ac-gfp) and HD-73Φ(cry1Ac-gfp)3534 were produced during asymmetric septum formation. Surprisingly, the Cry1Ac-GFP fusion protein showed polarity and was located near the septa in both strains. There was no significant difference between Cry1Ac-GFP and Cry1Ac in their toxicity to Plutella xylostella larvae.