Abstract
One possible strategy to delay the selection of resistant insect populations is the pyramiding of multiple resistance genes into a single cultivar. However, the transformation of most major crops remains prohibitively expensive if a large number of transgene combinations are to be evaluated. Arabidopsis thaliana is a potentially good plant for such preliminary evaluations. We determined that four major agricultural pests, Spodoptera exigua, Helicoverpa zea, Pseudoplusia includens, and Heliothis virescens grew as well when feeding on ‘Landsberg Erecta’ Arabidopsis as they did on plants of ‘Cobb’ soybean. Landsberg Erecta was then transformed with either a synthetic Bacillus thuringiensis cryIA(c) gene, or the cowpea trypsin inhibitor gene. Transformed plants were crossed to produce plants transgenic for both genes. Following quantification of transgene expression, the four caterpillar species were allowed to feed on wild-type plants, plants expressing either cryIA(c) or the cowpea trypsin inhibitor gene, or plants expressing both. Both genes reduced growth of the species tested, but cryIA(c) was more effective in controlling caterpillar growth than the cowpea trypsin inhibitor gene. The resistance of plants with both transgenes was lower than that of plants expressing the cryIA(c) gene alone, but higher than that of plants expressing the only the CpTI gene. This could be due to a lower concentration of Cry protein in the hemizygous F1 plants. Thus, if the cowpea trypsin inhibitor had any potentiation effect on cryIA(c), this effect was less than the cryIA(c) copy number effect. Alternatively, expression of the trypsin inhibitor gene could be antagonistic to the function of the cryIA(c) gene. Either way, these results suggest that the combined use of these two genes may not be effective.
Similar content being viewed by others
References
Adang MJ: Bacillus thuringiensis insecticidal crystal proteins: gene structure, action, and utilization. In: Maramorosch K (ed) Biotechnology for Biological Control of Pests and Vectors, pp. 3–24. CRC Press, Boca Raton, FL (1991).
Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K: Current Protocols in Molecular Biology: A Laboratory Manual. Greene Publishing Associates/Wiley-Interscience, New York (1989).
Beach RM, Todd JW: Resistance of the soybean breeding line GatIR81–296 to foliar feeding by the three Spodoptera sp. J Agric Entomol 4: 193–199 (1987).
Boulter D: Insect pest control by copying nature using genetically engineered crops. Phytochemistry. 34: 1453–1466 (1993).
Brattsten L: Bioengineering of crop plants and resistant biotype evolution in insects-counteracting coevolution. Arch Insect Biochem Physiol 17: 253–267 (1991).
Broadway RM: Tryptic inhibitory activity in wild and cultivated crucifers. Phytochemistry 28: 755–758 (1989).
Broadway RM: Are insects resistant to plant proteinase inhibitors? J Insect Physiol 41: 107–116 (1995).
Broadway RM, Colvin AA: Influence of cabbage proteinase inhibitors in situ on the growth of larval Trichoplusia ni and Pieris rapae. J Chem Ecol 18: 1009–1023 (1992).
Carozzi NB, Warren GW, Desai N, Jayne SM, Lotstein R, Rice DA, Evola S, Koziel MG: Expression of a chimeric CaMV 35S Bacillus thuringiensis insecticidal protein gene in transgenic tobacco. Plant Mol Biol 20: 539–548 (1992).
Doyle JJ, Doyle JL: Isolation of plant DNA from fresh tissue. Focus. 12: 13–15 (1990).
Gatehouse AMR, Boulter D, Hilder VA: Potential of plantderived genes in the genetic manipulation of crops for insect resistance. In: Gatehouse AMR, Hilder VA, Boulter D (eds) Plant Genetic Manipulation for Crop Protection, pp 155–181. Redwood Press, Melcksham (1992).
Gatehouse AMR, Shi Y, Powell KS, Brough C, Hilder VA, Hamilton WDO, Newell CA, Merryweather A, Boulter D, Gatehouse JA: Approaches to insect resistance using transgenic plants. Phil Trans R Soc Lond B 342: 279–286 (1993).
Geiger J, Fritz G: Determination of trypsin inhibition. In: Bergmeyer HU, Bergmeyer J, Grass M (eds) Methods of Enzymatic Analysis, pp. 121–126. VCH Publishers, Deerfield Beach, FL (1983).
Gibson DM, Gallo LG, Krasnoff SB, Ketchum REB: Increased efficacy of Bacillus thuringiensis subsp. kurstaki in combination with tannic acid. J Econ Entomol 88: 270–277 (1995).
Hilder VA, Gatehouse AMR, Sheerman SE, Barker RF, Boulter D: A novel mechanism of insect resistance engineered into tobacco. Nature. 330: 160–163 (1987).
Hoffman MP, Zalom FG, Wilson LT, Smilanick JM, Malyj LD, Kiser J, Hilder VA, Barnes W: Field evaluation of transgenic tobacco containing genes encoding Bacillus thuringiensis d-endotoxin or cowpea trypsin inhibitor: efficacy against Helicoverpa zea. J Econ Entomol 85: 2516–2522 (1992).
Hua X-J, Chen X-B, Ma W-C: Transgenic tobacco plants by cotransformation with proteinase inhibitor II and d-endotoxin genes. Chin Sci Bull 38: 1561–1566 (1993).
Johansen C: Principles of insect control. In: Pfadt R (ed) Fundementals of Applied Entomology, pp. 162–178. McMillan, New York (1985).
MacIntosh SC, Kishore GM, Perlak FJ, Marrone PG, Stone TB, Sims SR, Fuchs RL: Potentiation of Bacillus thuringiensis insecticidal activity by serine proteaase inhibitors. J Agric Food Chem 38: 1145–1152 (1990).
McGaughey WH: Problems of insect resistance to Bacillus thuringiensis. Agric Econ Envir 49: 95–102 (1994).
Murashige T, Skoog F: A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15: 473–497 (1962).
Murray EE, Roucheleau TR, Eberle M, Stock C, Sekar V, Adang MJ: Analysis of unstable RNA transcripts of insecticidal crystal protein genes of Bacillus thuringiensis in transgenic and electroporated protoplasts. Plant Mol Biol 16: 1035–1060 (1991).
Navon A, Hare JD, Federici BA: Interactions among Heliothis virescens larvae, cotton condensed tannin and the cryIA(c) δ-endotoxin of Bacillus thuringiensis. J Chem Ecol 19: 2485–2499 (1993).
Pratt LH, McCurdy DW, Shimazaki Y, Cordonnier M-M: Immunodetection of phytochrome: immunocytochemistry, immunoblotting, and immunoquantitation. In: Linskens HF, Jackson JF (eds) Modern Methods in Plant Analysis, pp. 50–74. Springer-Verlag, New York (1986).
Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1989).
Singset C, Adang MJ, Lynch RE, Anderson WF, Wang A, Cardineau G, Ozias-Akins P: Expression of Bacillus thuringiensis cryIA(c) gene in transgenic peanut and its efficacy against lesser cornstalk borer. Transgen Res. in press (1997).
Stewart CN, Adang MJ, All JN, Boerma R, Cardineau G, Tucker D, Parrott WA: Genetic transformation, recovery, and characterization of soybean (Glycine max (L.) Merrill) transgenic for a synthetic Bacillus thuringiensis cryIA(c) gene. Plant Physiol 112: 121–129 (1997).
Stewart CN, Adang MJ, All JN, Raymer PL, Ramachandran S, Parrott WA: Insect control and dosage effects in transgenic canola, Brassica napus L. (Brassicaceae), containing a synthetic Bacillus thuringiensis cryIA(c) gene. Plant Physiol 112: 115–120 (1997).
Tabashnik BE: Evolution of resistance to Bacillus thuringiensis. Annu Rev Entomol 39: 47–79 (1994).
Tabashnik BE, Cushing NL, Finson N, Johnson NW: Field development of resistance to Bacillus thuringiensis in diamondback moth. J Econ Entomol 83: 1671–1676 (1990).
Tabashnik BE, Finson N, Johnson MW: Two protease inhibitors fail to synergize Bacillus thuringiensis in diamondback moth (Lepidoptera: Plutellidae). J Econ Entomol 85: 2082–2087 (1992).
Tabashnik BE, Finson N, Johnson MW, Heckel DG: Crossresistance to Bacillus thuringiensis toxin CryIF in the diamondback moth. Appl Environ Microbiol 60: 4627–4629 (1994).
Vaeck M, Reynaerts A, Höfte H, Jansens S, De Beuckeleer M, Dean C, Zabeau M, Van Montagu M, Leemans J: Transgenic plants protected from insect attack. Nature 328: 33–37 (1987).
Valvekens D, Van Montagu M, Van Lijsebettens M: Agrobacterium tumefaciens-mediated transformation of Arabidopsis thaliana root explants by using kanamycin selection. Proc Natl Acad Sci USA 85: 5536–5540 (1988).
Van Rie J: Insect control with transgenic plants: resistance proof? Trends Biotechnol 9: 177–179 (1991).
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Santos, M.O., Adang, M.J., All, J.N. et al. Testing transgenes for insect resistance using Arabidopsis. Molecular Breeding 3, 183–194 (1997). https://doi.org/10.1023/A:1009603101549
Issue Date:
DOI: https://doi.org/10.1023/A:1009603101549