Two different Bacillus thuringiensis toxin genes confer resistance to beet armyworm (Spodoptera exigua Hübner) in transgenic Bt-shallots (Allium cepa L.)

S.J. Zheng Sijun, B. Henken, R.A. de Maagd, A. Purwito, F.A. Krens, C. Kik

Research output: Contribution to journalArticleAcademicpeer-review

23 Citations (Scopus)

Abstract

Agrobacterium-mediated genetic transformation was applied to produce beet armyworm (Spodoptera exigua Hübner) resistant tropical shallots (Allium cepa L. group Aggregatum). A cry1Ca or a H04 hybrid gene from Bacillus thuringiensis, driven by the chrysanthemum ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit (Rubisco SSU) promoter, along with the hygromycin phosphotransferase gene (hpt) driven by the CaMV 35S promoter, was employed for genetic transformation. An average transformation frequency of 3.68% was obtained from two shallot cultivars, Tropix and Kuning. After transfer of the in vitro plants to the greenhouse 69% of the cry1Ca and 39% of the H04 transgenic shallots survived the first half year. After one year of cultivation in the greenhouse the remaining cry1Ca and H04 transgenic plants grew vigorously and had a normal bulb formation, although the cry1Ca transgenic plants (and controls) had darker green leaves compared to their H04 counterparts. Standard PCR, adaptor ligation PCR and Southern analyses confirmed the integration of T-DNA into the shallot genome. Northern blot and ELISA analyses revealed expression of the cry1Ca or H04 gene in the transgenic plants. The amount of Cry1Ca expressed in transgenic plants was higher than the expression levels of H04 (0.39 vs. 0.16% of the total soluble leaf proteins, respectively). There was a good correlation between protein expression and beet armyworm resistance. Cry1Ca or H04 gene expression of at least 0.22 or 0.08% of the total soluble protein in shallot leaves was sufficient to give a complete resistance against beet armyworm. This confirms earlier observations that the H04 toxin is more toxic to S. exigua than the Cry1Ca toxin. The results from this study suggest that the cry1Ca and H04 transgenic shallots developed could be used for introducing resistance to beet armyworm in (sub) tropical shallot
Original languageEnglish
Pages (from-to)261-272
JournalTransgenic Research
Volume14
Issue number3
DOIs
Publication statusPublished - 2005

Fingerprint

Shallots
shallots
Spodoptera
Bacillus thuringiensis
Beta vulgaris
Spodoptera exigua
Onions
Allium cepa
toxins
genetically modified organisms
Genetically Modified Plants
transgenic plants
Genes
genes
Genetic Transformation
hygromycin-B kinase
genetic transformation
promoter regions
Chrysanthemum
Ribulose-Bisphosphate Carboxylase

Keywords

  • tumefaciens-mediated transformation
  • synthetic sex-pheromone
  • delta-endotoxin
  • communication disruption
  • plant-regeneration
  • domain iii
  • agrobacterium
  • lepidoptera
  • noctuidae
  • dna

Cite this

@article{a7a3196841074e2f99b039c087340b3b,
title = "Two different Bacillus thuringiensis toxin genes confer resistance to beet armyworm (Spodoptera exigua H{\"u}bner) in transgenic Bt-shallots (Allium cepa L.)",
abstract = "Agrobacterium-mediated genetic transformation was applied to produce beet armyworm (Spodoptera exigua H{\"u}bner) resistant tropical shallots (Allium cepa L. group Aggregatum). A cry1Ca or a H04 hybrid gene from Bacillus thuringiensis, driven by the chrysanthemum ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit (Rubisco SSU) promoter, along with the hygromycin phosphotransferase gene (hpt) driven by the CaMV 35S promoter, was employed for genetic transformation. An average transformation frequency of 3.68{\%} was obtained from two shallot cultivars, Tropix and Kuning. After transfer of the in vitro plants to the greenhouse 69{\%} of the cry1Ca and 39{\%} of the H04 transgenic shallots survived the first half year. After one year of cultivation in the greenhouse the remaining cry1Ca and H04 transgenic plants grew vigorously and had a normal bulb formation, although the cry1Ca transgenic plants (and controls) had darker green leaves compared to their H04 counterparts. Standard PCR, adaptor ligation PCR and Southern analyses confirmed the integration of T-DNA into the shallot genome. Northern blot and ELISA analyses revealed expression of the cry1Ca or H04 gene in the transgenic plants. The amount of Cry1Ca expressed in transgenic plants was higher than the expression levels of H04 (0.39 vs. 0.16{\%} of the total soluble leaf proteins, respectively). There was a good correlation between protein expression and beet armyworm resistance. Cry1Ca or H04 gene expression of at least 0.22 or 0.08{\%} of the total soluble protein in shallot leaves was sufficient to give a complete resistance against beet armyworm. This confirms earlier observations that the H04 toxin is more toxic to S. exigua than the Cry1Ca toxin. The results from this study suggest that the cry1Ca and H04 transgenic shallots developed could be used for introducing resistance to beet armyworm in (sub) tropical shallot",
keywords = "tumefaciens-mediated transformation, synthetic sex-pheromone, delta-endotoxin, communication disruption, plant-regeneration, domain iii, agrobacterium, lepidoptera, noctuidae, dna",
author = "{Zheng Sijun}, S.J. and B. Henken and {de Maagd}, R.A. and A. Purwito and F.A. Krens and C. Kik",
note = "2680",
year = "2005",
doi = "10.1007/s11248-005-0109-2",
language = "English",
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pages = "261--272",
journal = "Transgenic Research",
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}

Two different Bacillus thuringiensis toxin genes confer resistance to beet armyworm (Spodoptera exigua Hübner) in transgenic Bt-shallots (Allium cepa L.). / Zheng Sijun, S.J.; Henken, B.; de Maagd, R.A.; Purwito, A.; Krens, F.A.; Kik, C.

In: Transgenic Research, Vol. 14, No. 3, 2005, p. 261-272.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Two different Bacillus thuringiensis toxin genes confer resistance to beet armyworm (Spodoptera exigua Hübner) in transgenic Bt-shallots (Allium cepa L.)

AU - Zheng Sijun, S.J.

AU - Henken, B.

AU - de Maagd, R.A.

AU - Purwito, A.

AU - Krens, F.A.

AU - Kik, C.

N1 - 2680

PY - 2005

Y1 - 2005

N2 - Agrobacterium-mediated genetic transformation was applied to produce beet armyworm (Spodoptera exigua Hübner) resistant tropical shallots (Allium cepa L. group Aggregatum). A cry1Ca or a H04 hybrid gene from Bacillus thuringiensis, driven by the chrysanthemum ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit (Rubisco SSU) promoter, along with the hygromycin phosphotransferase gene (hpt) driven by the CaMV 35S promoter, was employed for genetic transformation. An average transformation frequency of 3.68% was obtained from two shallot cultivars, Tropix and Kuning. After transfer of the in vitro plants to the greenhouse 69% of the cry1Ca and 39% of the H04 transgenic shallots survived the first half year. After one year of cultivation in the greenhouse the remaining cry1Ca and H04 transgenic plants grew vigorously and had a normal bulb formation, although the cry1Ca transgenic plants (and controls) had darker green leaves compared to their H04 counterparts. Standard PCR, adaptor ligation PCR and Southern analyses confirmed the integration of T-DNA into the shallot genome. Northern blot and ELISA analyses revealed expression of the cry1Ca or H04 gene in the transgenic plants. The amount of Cry1Ca expressed in transgenic plants was higher than the expression levels of H04 (0.39 vs. 0.16% of the total soluble leaf proteins, respectively). There was a good correlation between protein expression and beet armyworm resistance. Cry1Ca or H04 gene expression of at least 0.22 or 0.08% of the total soluble protein in shallot leaves was sufficient to give a complete resistance against beet armyworm. This confirms earlier observations that the H04 toxin is more toxic to S. exigua than the Cry1Ca toxin. The results from this study suggest that the cry1Ca and H04 transgenic shallots developed could be used for introducing resistance to beet armyworm in (sub) tropical shallot

AB - Agrobacterium-mediated genetic transformation was applied to produce beet armyworm (Spodoptera exigua Hübner) resistant tropical shallots (Allium cepa L. group Aggregatum). A cry1Ca or a H04 hybrid gene from Bacillus thuringiensis, driven by the chrysanthemum ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit (Rubisco SSU) promoter, along with the hygromycin phosphotransferase gene (hpt) driven by the CaMV 35S promoter, was employed for genetic transformation. An average transformation frequency of 3.68% was obtained from two shallot cultivars, Tropix and Kuning. After transfer of the in vitro plants to the greenhouse 69% of the cry1Ca and 39% of the H04 transgenic shallots survived the first half year. After one year of cultivation in the greenhouse the remaining cry1Ca and H04 transgenic plants grew vigorously and had a normal bulb formation, although the cry1Ca transgenic plants (and controls) had darker green leaves compared to their H04 counterparts. Standard PCR, adaptor ligation PCR and Southern analyses confirmed the integration of T-DNA into the shallot genome. Northern blot and ELISA analyses revealed expression of the cry1Ca or H04 gene in the transgenic plants. The amount of Cry1Ca expressed in transgenic plants was higher than the expression levels of H04 (0.39 vs. 0.16% of the total soluble leaf proteins, respectively). There was a good correlation between protein expression and beet armyworm resistance. Cry1Ca or H04 gene expression of at least 0.22 or 0.08% of the total soluble protein in shallot leaves was sufficient to give a complete resistance against beet armyworm. This confirms earlier observations that the H04 toxin is more toxic to S. exigua than the Cry1Ca toxin. The results from this study suggest that the cry1Ca and H04 transgenic shallots developed could be used for introducing resistance to beet armyworm in (sub) tropical shallot

KW - tumefaciens-mediated transformation

KW - synthetic sex-pheromone

KW - delta-endotoxin

KW - communication disruption

KW - plant-regeneration

KW - domain iii

KW - agrobacterium

KW - lepidoptera

KW - noctuidae

KW - dna

U2 - 10.1007/s11248-005-0109-2

DO - 10.1007/s11248-005-0109-2

M3 - Article

VL - 14

SP - 261

EP - 272

JO - Transgenic Research

JF - Transgenic Research

SN - 0962-8819

IS - 3

ER -