Open Access Open Access  Restricted Access Subscription or Fee Access
Cover Image

Expression of Insecticidal Toxin Coded by Modified Full-Length and Truncated Bt-cry1Ac Gene in Transgenic Tomato for Assessment of Their Stability and Efficacy against Target Insects.

Bhupendra Koul, Devendra V. Amla, Indraneel Sanyal


The work is structured into four different parts. The first part is dedicated to computational analysis of six variants of Bt-cry1A genes to generate an idea of their integrity and stability. The second part includes optimization of Agrobacterium-mediated tomato transformation procedure using vegetative leaf disc-explants with p35SGUS-INT and validation of the procedure using five different genes of diverse origin. The third part deals with a comparative analysis of constitutive promoter DECaMV35S and synthetic promoter Pcec (complete expression cassette) with defined regulatory sequences to enhance the expression and efficacy of Bt-cry1Ac gene against Helicoverpa armigera in transgenic tomato. Finally, the fourth part discusses molecular characterization and comparative expression analysis of transgenic tomato transformed with modified full-length and truncated version of cry1Ac genes. It also includes the performance and efficacy of truncated cry1Ab gene against Helicoverpa armigera and Spodoptera litura, in transgenic tomato. The higher level of Cry1Ab toxin (≈ 0.47 ± 0.01% TSP) did not show any detrimental effect on in vitro regeneration, plant development and efficacy against target pests in transgenic line Ab25E, contrary to the  earlier reports with Cry1Ac toxin. The full-length cry1Ac gene can be redesigned for higher expression and performance in dicots.


Transgenic tomato, Vegetative leaf, Transformation efficiency, Expression analysis, Helicoverpa armigera

Full Text:



E.E. Murray, J. Lotzer, M. Eberle. Codon usage plant genes. Nucl. Acids Res 1989, 17, 477–498.

F.J. Perlak, R.L. Fuchs, D.A. Dean, S.L. McPherson, D.A. Fischhoff .Modification of the coding sequences enhances plant expression of insect control protein genes. Proc. Natl. Acad. Sci. USA 1991, 88, 3324–3328.

M.G. Koziel, G.L. Beland, C. Bowman, N.B. Carozzi, R. Crenshaw, L. Crossland, J. Dawson, N. Desai, M. Hill, S. Kadwell, K. Launis, K. Lewis, D. Maddox, K. McPherson, M.R. Meghji, E. Merlin, R. Rhodes, G.W. Warren, M.Wright, S.V. Evola. Field performance of elite transgenic maize plants expressing an insecticidal protein derived from Bacillus thuringiensis. Bio/Tech. 1993,11,194–200.

D.A. Fischhoff , K.S. Bowdish, F.J. Perlak, P.G. Marrone, S.M. McCormick, J.G. Niedermeyer, D.A. Dean, K. Kusano-Kretzmer, E.J. Mayer, D.E. Rochester, S.G. Rogers, R.T. Fraley Insect tolerant transgenic tomato plants. Bio/Tech. 1987, 5, 807–813.

R.T. Roush Bt transgenic crops: just another pretty insecticide or a chance for a new start in resistance management. Pestc. Sci. 1997, 51, 328–334.

J.J. Estruch, N.B. Carozzi, N. Desai, N.B. Duck, G.W. Warren, M.G. Koziel. Transgenic plants: an emerging approach to pest control. Nat. Biotech. 1997, 15, 137–141.

J. VanRie Bacillus thuringiensis and its use in transgenic insect control technologies. Int. J. Med. Microbiol. 2000, 290, 463–469.

S.H. Diehn., E.J. De Rocher, P.J. Green. In Problems that can limit the expression of foreign genes in plants: lessons to be learned from B.t.-toxin genes. J.K. Setlow, Ed.; Genetic Engineering: Principles and Methods: Plenum Press, New York, 1996, Vol. 18, pp 83–99.

E. J. D. Rocher, T. C. Vargo-Gogola, S. H. Diehn, P. J. Green. Direct evidence for rapid degradation of Bacillus thuringiensis toxin mRNA as a cause of poor expression in plants. Plant Physiol. 1998, 117, 1445–1461.

P. Rawat, A.K.Singh, K. Ray, B. Chaudhary, S. Kumar, T. Gautam, S. Kanoria, G. Kaur, P. Kumar, D. Pental, P.K. Burma Detrimental effect of expression of Bt endotoxin Cry1Ac on in vitro regeneration, in vivo growth and development of tobacco and cotton transgenics. J. Biosci. 2011, 36, 363–376.

H. Kumar, V. Kumar Tomato expressing Cry1A(b) insecticidal protein from Bacillus thuringiensis protected against tomato fruit borer, Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae) damage in the laboratory, greenhouse and field. Crop Prot. 2004, 23,135–139.

S. Srivastava. Ph.D. Thesis, Expression and performance of modified Bacillus thuringiensis insecticidal cry1A genes in transgenic tomato for insect resistance, University of Lucknow, Lucknow, India, 2007.

C. James. Global status of commercialized biotech/GM crops. ISAAA Brief No. 44. ISAAA: Ithaca NY, 2012.

P. Bhatia, N. Ashwath, T. Senaratna, D. Midmore Tissue Culture Studies of Tomato (Lycopersicon esculentum). Plant Cell Tiss. Org. Cult. 2004, 78, 1–21.

D. Liénard, C. Sourrouille, V. Gomord, L. Faye. Pharming and transgenic plants. Biotechnol. Ann. Rev. 2007, 13,115–147.

C. Halpin. Gene stacking in transgenic plants-the challenge for 21st century plant biotechnology. Plant Biotechnol. J. 2005, 3,141–155.

M.J. Battraw, T.C. Hall. Histochemical analysis of CaMV35S promoter β-glucuronidase gene expression in transgenic rice plants. Plant Mol. Biol. 1990, 15, 527–538.

P.N. Benfey, L. Ren, N.H. Chua. The CaMV35S enhancer contains at least two domains, which can confer different developmental and tissue-specific expression patterns. EMBO. J. 1989, 8, 2195–2202.

I.S. Curtis, J.B. Power, P. Hedden, A. Phillip, K.C. Lowe, D.A.Ward, M.R. Davey. Transformation and characterization of transgenic plants of Solanum dulcamara L. –incidence of transgene silencing. Ann. Bot. 2000, 86, 63–71.

K. Gogoi. Synthesis and biophysical studies of PNA and chimeric PNA-DNA antisense oligomers with five atom linkages. Int. Archives Sci. Technol., 2013, 13(1), 7-13.

S.L.A. Hobbs, T.D.Warkentin, C.M.O. DeLong. Transgene copy number can be positively or negatively associated with transgene expression. Plant Mol. Biol. 1993, 21, 17–26.

W.Z. Guo, J. Sun, Y.F. Guo, T.Z. Zhang Investigation of different dosage of inserted Bt genes and their insect-resistance in transgenic Bt cotton. Acta Genet. Sin. 2001, 28, 668–676.

C.K. Rao. Transgenic Bt Technology: 3. Expression of Transgenes. Available from news/ukshowlib.phtml?uid¼9304, 2005.

J. Singh. Effect of heavy metals and sewage on seed germination and plant growth. Int. Archives Sci. Technol., 2006, 6(1), 1-4.

B. Koul, R. Yadav, I. Sanyal, S. Sawant, V. Sharma, D.V. Amla. Cis-acting motifs in artificially synthesized expression cassette leads to enhanced transgene expression in tomato (Solanum lycopersicum L.) Plant Physiol. & Biochem. 2012, 61, 131-141.

J. Singh. Determination of DTPA extractable heavy metals from sewage irrigated fields and plants. J. Integr. Sci. Technol., 2013, 1(1), 36-40.

ISSN 2347 – 8853

Indexed in: