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Scientia Silvae Sinicae ›› 2005, Vol. 41 ›› Issue (4): 118-122.doi: 10.11707/j.1001-7488.20050420

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Construction of A Pine Needle Symbiotic Engineered Bacterium Exhibiting the Insecticidal Activity to Dendrolimus punctatus

Zhao Tonghai1,Xu Jing2,Xu Hongmei3,Zhang Qingwen2,Chen Jingyuan3   

  1. 1.Institute of Forest Ecology, Environment and Protection, CAF Beijing 100091; 2. Department of Entomology, China Agricultural University Beijing10094; 3.Institute of Forest Protection, Hubei Academy of Forestry Wuhan430079
  • Received:2003-09-18 Revised:1900-01-01 Online:2005-07-25 Published:2005-07-25

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Abstract:

Bt cryIAc gene from Bacillus thuringiensis subsp. Kurstaki was introduced into the chromosome of pine symbiotic bacterium Bacillus cereus (Bc752) by an integrative vector pEG601. The vector contains a strong vegetative promoter, the cryIAc gene (identified as a 7.4 kb SphI-NruI fragment of DNA from B. thuringiensis HD-73), the tetracycline resistance gene (tetr) and an 8.0 kb EcoRI-NcoI fragment of chromosomal DNA from Bc752 to allow for homologous recombinant between the vector and the bacterial chromosome. Transformation of Bc752 with plasmid pEG601 by electroporation resulted in the engineered symbiotic bacteria. Insertion of the vector DNA into the chromosome was demonstrated by analysis of DNA restriction enzymes and PCR amplification. Recombination strains containing Bt cryIAc gene produced the 133ku protoxin which could be detected by SDS-PAGE, ELISA, electronmicroscope observation and insect bioassays.

Key words: Bacillus thuringiensis(Bt), insecticidal protein, pine needle symbiotic bacteria, engineered bacterium, Dendrolimus punctatus