title: In vivo fluorescence observation of parasporal inclusion formation in Bacillus thuringiensis.
abstract: 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(-), and the resulting strain was named HD-73(-)(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(-)(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.
authors.affiliation: State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Yang
authors.forename: Rong
authors.initials: R
authors.lastname: Rong
authors.forename: FuPing
authors.initials: F
authors.lastname: Song
authors.forename: ChangPo
authors.initials: C
authors.lastname: Sun
authors.forename: Juan
authors.initials: J
authors.lastname: Wei
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.forename: DaFang
authors.initials: D
authors.lastname: Huang
mesh_terms: D000818:Animals; D001413:Bacillus thuringiensis / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004731:Endotoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002479:Inclusion Bodies / Q000737:chemistry / Q000378:metabolism; D007306:Insecticides / Q000737:chemistry / Q000494:pharmacology; D007814:Larva / Q000187:drug effects; D009036:Moths / Q000187:drug effects / Q000254:growth & development; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013171:Spores, Bacterial / Q000737:chemistry / Q000378:metabolism
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D007306:Insecticides; D011993:Recombinant Fusion Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis