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Cloning and Characterization of Two Novel Crystal Protein Genes, cry54Aa1 and cry30Fa1, from Bacillus thuringiensis Strain BtMC28

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Abstract

Bacillus thuringiensis strain BtMC28 was isolated from the soil sample in China. Two novel crystal protein genes were found by using the PCR-RFLP method. Moreover, the full-length sequences of two novel genes were obtained by a single oligonucleotide nested (SON)-PCR upstream and downstream strategy. Sequence analysis revealed that one gene encoded a polypeptide of 673 amino acid residues with a molecular mass of 76.3 kDa, 38% identical to Cry10Aa, and the other encoded a polypeptide of 687 amino acid residues with a molecular mass of 77.1 kDa, 74% identical to Cry30Aa. These two novel crystal protein genes were designated as cry54Aa1 and cry30Fa1 by Bt Insecticidal Crystal Proteins Nomenclature Committee, respectively. The Cry54Aa1 and Cry30Fa1 proteins retained five conserved regions commonly found in the existing Cry proteins. Cry54Aa1 protein exhibited insecticidal activities against Laphygma exigua (Lepidoptera), Helicoverpa armigera (Lepidoptera), and Aedes aegypti (Diptera) when its encoding gene was expressed in an Escherichia coli host strain.

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References

  1. Antal ZC, Rascle M, Fevre C, Bruel C (2004) Single oligonucleotide nested PCR: a rapid method for the isolation of genes and their flanking regions from expressed sequence tags. Curr Genet 46:240–246

    Article  PubMed  CAS  Google Scholar 

  2. Boonserm P, Davis P, Ellar DJ, Li J (2005) Crystal structure of the mosquito-larvicidal toxin Cry4Ba and its biological implications. J Mol Biol 348:363–382

    Article  PubMed  CAS  Google Scholar 

  3. Galitsky N, Cody V, Wojtczak A, Ghosh D, Luft JR, Pangborn W, English L (2001) Structure of the insecticidal bacterial δ-endotoxin Cry3Bb1 of Bacillus thuringiensis. Acta Crystallogr D Biol Crystallogr 57:1101 1099

    Article  PubMed  CAS  Google Scholar 

  4. Georghiou GP, Wirth MC (1997) Influence of exposure to single versus multiple toxins of Bacillus thuringiensis subsp. israelensis on development of resistance in the mosquito Culex quinquefasciatus (Diptera: Culicidae). Appl Environ Microbiol 63:1095–1101

    PubMed  CAS  Google Scholar 

  5. Grochulski P, Masson L, Borisova S, Pusztai-Carey M, Schwartz J, Brousseau R, Cygler M (1995) Bacillus thuringiensis CryIA(a) insecticidal toxin: crystal structure and channel formation. J Mol Biol 254:447–464

    Article  PubMed  CAS  Google Scholar 

  6. Hofte HE, Whiteley HR (1989) Insecticidal crystal proteins of Bacillus thuringiensis. Microbiol Rev 53:242–255

    PubMed  CAS  Google Scholar 

  7. Ibarra JE, del Rincón MC, Ordúz S, Noriega D, Benintende G, Monnerat R, Regis L, de Oliverira CMF, Lanz H, Rodriguez MH, Sánchez J, Peíía G, Bravo A (2003) Diversity of Bacillus thuringienisis strains from Latin America with insecticidal activity against different mosquito species. Appl Environ Microbiol 69:5269–5274

    Article  PubMed  CAS  Google Scholar 

  8. Kwo WS, Chak KF (1996) Identification of novel cry-type genes from Bacillus thuringiensis strains on the basis of restriction fragment length polymorphism of the PCR-amplified DNA. Appl Environ Microbiol 62:1369–1377

    Google Scholar 

  9. Li JD, Carroll J, Ellar DJ (1991) Crystal structure of insecticidal δ-endotoxin from Bacillus thuringiensis at 2.5 A resolution. Nature 353:815–821

    Article  PubMed  CAS  Google Scholar 

  10. Morse RJ, Yamamoto T, Stroud RM (2001) Structure of Cry2Aa suggests an unexpected receptor binding epitope. Structure 9:409 407

    Article  PubMed  CAS  Google Scholar 

  11. Narva KE, Payne JM, Schwab GE, Hickle LA, Galasan T, Sick AJ (1991) Novel Bacillus thuringiensis microbes active against nematodes, and genes encoding novel nematodes- active toxins cloned from Bacillus thuringiensis isolates. European Patent Office, EP 0462721

  12. Sambrook JE, Fritsch F, Maaiatis T (2002) A laboratory of molecular cloning, 3rd edn. Cold Spring Harbor Laboratory Press, New York

    Google Scholar 

  13. Schnepf HE, Whiteley HR (1981) Cloning and expression of the Bacillus thuringiensis crystal protein gene in Escherichia coli. Proc Natl Acad Sci USA 78:2893–2897

    Article  PubMed  CAS  Google Scholar 

  14. Schnepf HE, Crickmore N, Van Rie J, Lereclus D, Baum J, Feitelson J, Zeigler DR, Dean DH (1998) Bacillus thuringiensis and its pesticidal crystal proteins. Microbiol Mol Biol Rev 62:775–806

    PubMed  CAS  Google Scholar 

  15. Servant F, Bru C, Carrère S, Courcelle E, Gouzy J, Peyruc D, Kahn D (2002) ProDom: automated clustering of homologous domains. Brief Bioinform 3:246–251

    Article  PubMed  CAS  Google Scholar 

  16. Song FP, Zhang J, Chen ZY, Huang DF (1998) Establishment of PCR-RFLP identification system of cry genes from Bacillus thuringiensis. Agric Sci China 31:13–18

    CAS  Google Scholar 

  17. Song FP, Zhang J, Gu AX, Wu Y, Han LL, He KL, Chen ZY, Yao J, Hu YQ, Li GX, Huang DF (2003) Identification of cry1I-type genes from Bacillus thuringiensis strains and characterization of a novel cry1I-type gene. Appl Environ Microbiol 69:5207–5211

    Article  PubMed  CAS  Google Scholar 

  18. Su XD (2005) Isolation of B. thuringiensis strains and identification of their cry genes. M.S. Thesis, Agricultural University of Hebei, Baoding

  19. Tabashnik BE, Cushing NL, Finson N, Johnson MW (1990) Field development of resistance to Bacillus thuringiensis in diamondback moth (Lepidoptera: Plutellidae). J Econ Entomol 83:1671–1676

    Google Scholar 

  20. Thorne L, Garduno F, Thompson T, Decker D, Zounes M, Wild M, Walfield M, Pollock TJ (1986) Structural similarity between the lepidoptera- and diptera-specific insecticidal endotoxin genes of Bacillus thuringiensis subsp. “kurstaki” and “israelensis”. J Bacteriol 166:801–811

    PubMed  CAS  Google Scholar 

  21. Yu H, Zhang J, Huang DF, Gao JG, Song FP (2006) Characterization of Bacillus thuringiensis strain Bt185 toxic to the Asian Cockchafer: Holotrichia parallela. Curr Microbiol 53:13–17

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

We are grateful to Professor Jie Zhang and Fuping Song (Institute of Plant Protection, Chinese Academy of Agricultural Sciences) for helpful suggestions. This study was supported by a grant from the National High Technology Research and Development Program of China (863 Program) (No. 2006AA02Z189).

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Authors and Affiliations

  1. Rice Research Institute, Sichuan Agricultural University, Yaan, China

    Furong Tan, Jun Zhu, Jie Tang, Shiquan Wang, Aiping Zheng & Ping Li

  2. Biotechnology Research Center, Shanghai Academy of Agriculture Sciences, Shanghai, China

    Furong Tan & Xueming Tang

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  1. Furong Tan
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  2. Jun Zhu
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  3. Jie Tang
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  4. Xueming Tang
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  5. Shiquan Wang
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  6. Aiping Zheng
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  7. Ping Li
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Correspondence to Aiping Zheng or Ping Li.

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The authors, Furong Tan and Jun Zhu contributed equally to this work.

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Tan, F., Zhu, J., Tang, J. et al. Cloning and Characterization of Two Novel Crystal Protein Genes, cry54Aa1 and cry30Fa1, from Bacillus thuringiensis Strain BtMC28. Curr Microbiol 58, 654–659 (2009). https://doi.org/10.1007/s00284-009-9386-y

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  • DOI: https://doi.org/10.1007/s00284-009-9386-y

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