Agron. Sustain. Dev.
Volume 28, Number 4, October-December 2008
Page(s) 481 - 488
Published online 21 June 2008
Agron. Sustain. Dev. 28 (2008) 481-488
DOI: 10.1051/agro:2008025

Transgenic resistance of Bulgarian potato cultivars to the Colorado potato beetle based on Bt technology

Ivanka Kamenova1, Rossitza Batchvarova1, Stanislaw Flasinski2, Lidia Dimitrova3, Petya Christova1, Slavcho Slavov1, Atanas Atanassov1, Plamen Kalushkov4 and Wojciech Kaniewski5

1  AgroBioInstitute, 8 Dragan Tzankov Blvd., 1164 Sofia, Bulgaria
2  Monsanto, GG3220, 700 Chesterfield Parkway West, St. Louis, MO 63017, USA
3  Potato Laboratory, 2000 Samokov, Bulgaria
4  Institute of Zoology, Bulgarian Academy of Sciences, 1 Zar Osvoboditel, 1000 Sofia, Bulgaria
5  Adam Mickiewicz University, 85 Umultowska ul. 61-614 Poznan, Poland

Accepted 30 April 2008; published online 21 June 2008

Abstract - Colorado potato beetle, Leptinotarsa decemlineata Say, is the most destructive insect pest of potatoes. When the population of beetles is high, plants can be completely defoliated and commercial potato production is nearly impossible without control of the beetle. The beetles have shown a tremendous ability to develop resistance against insecticides. Previously, a biotechnology approach to control Colarado potato beetle based on the use of the synthetic Bt gene was developed. In this article, a transformation procedure for three commercial Bulgarian potato cultivars was developed and potentially commercial transgenic lines have been selected based on field resistance to Colorado potato beetles and yield. Plants were transformed with the Bacillus thuringiensis (Bt) cry3A gene using Agrobacterium-mediated transformation. 110 plants from the three cultivars were regenerated and tested by double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). The Cry3A protein accumulation varied across the transgenic lines, rating from very low to 71.5 $\mu $g/g fresh weight. 21 transgenic lines expressing the Cry3A protein at levels above 10 $\mu $g/g fresh weight were tested in two successive years in filed conditions at two different locations of the country. All transgenic lines compared with the controls, nontransgenic potatoes from the respective cultivar, were consistently protected from foliar damages from all developmental stages of the beetle. The comparison of all properties of the tested transgenic lines, including variety phenotypes and tuber yield, allowed the selection of the most promising 2–3 lines per cultivar. Selected lines produced tuber amounts 80–100% higher compared with the control, non-transgenic plants. Those lines were grown for mass propagation during the third year of field experiments. The presence of the transgene in these lines was confirmed with the use of primers specific to the transgene by polymerase chain reaction (PCR). Additionally, the results from the insect bioassay showed that these lines were highly resistant to insect feeding, leading to 100% of mortality of larval populations. In summary, we generated potentially commercial potato lines highly resistant to Colorado potato beetle using Bt technology that may have a profound impact on development of sustainable agriculture in Bulgaria. This is one of the several agriculture biotechnology products entirely developed and tested in Bulgaria.

Key words: potato / Bt Cry3A / Colorado potato beetle resistance / field tests / genetic transformation

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© INRA, EDP Sciences 2008