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Applied and Environmental Microbiology, November 2008, p. 6997-7001, Vol. 74, No. 22
0099-2240/08/$08.00+0     doi:10.1128/AEM.01346-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

New Strategy for Isolating Novel Nematicidal Crystal Protein Genes from Bacillus thuringiensis Strain YBT-1518{triangledown}

Suxia Guo,1 Mei Liu,1,2 Donghai Peng,1 Sisi Ji,1 Pengxia Wang,1 Ziniu Yu,1 and Ming Sun1*

State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology,1 College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, Hubei, People's Republic of China2

Received 16 June 2008/ Accepted 19 September 2008

We have developed a strategy for isolating cry genes from Bacillus thuringiensis. The key steps are the construction of a DNA library in an acrystalliferous B. thuringiensis host strain and screening for the formation of crystal through optical microscopy observation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analyses. By this method, three cry genes—cry55Aa1, cry6Aa2, and cry5Ba2—were cloned from rice-shaped crystals, producing B. thuringiensis YBT-1518, which consists of 54- and 45-kDa crystal proteins. cry55Aa1 encoded a 45-kDa protein, cry6Aa2 encoded a 54-kDa protein, and cry5Ba2 remained cryptic in strain YBT-1518, as shown by SDS-PAGE or microscopic observation. Proteins encoded by these three genes are all toxic to the root knot nematode Meloidogyne hapla. The two genes cry55Aa1 and cry6Aa2 were found to be located on a plasmid with a rather small size of 17.7 kb, designated pBMB0228.

* Corresponding author. Mailing address: State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, Hubei, People's Republic of China. Phone: 86-27-87283455. Fax: 86-27-87280670. E-mail: m98sun{at}mail.hzau.edu.cn

{triangledown} Published ahead of print on 26 September 2008.

Applied and Environmental Microbiology, November 2008, p. 6997-7001, Vol. 74, No. 22
0099-2240/08/$08.00+0     doi:10.1128/AEM.01346-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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