Agrobacterium-mediated genetic transformation of cauliflower with the NPK1 gene to enhance biotic and abiotic stress tolerance
Authorized Users Only
2011
Authors
Pavlović, SuzanaVinterhalter, Branka
Zdravković-Korać, Snežana
Adžić, Slađan
Đorđević, Mladen
Ugrinović, Milan
Mitić, Nevena
Conference object (Published version)
Metadata
Show full item recordAbstract
Cauliflower (Brassica oleracea var. botrytis) is an important vegetable plant grown worldwide that is impacted by biotic and abiotic stresses. Plant biotechnology could be an efficient approach to enhance stress tolerance. Mitogen-activated protein kinase (MAPK) cascades play a key role in plant growth and development as well as biotic and abiotic stress response. Thus, we have established an Agrobaterium tumefacens-mediated protocol for transformation of cauliflower (line KFRM4) using the binary vector pSHX004 with the mitogen-activated protein kinase kinase kinase (MAPKKK) gene from tobacco (NPK1) for stress tolerance and the phosphinothricin acetyltransferase (bar) selectable marker gene for herbicide resistance. The effect of acetosyringone (AS) on transformation efficiency was also investigated. Hypocotyls of 8 day-old seedlings, pre-cultivated on MS shoot induction medium (SIM) supplemented with 1.0 mg/l BA and 0.2 mg/l IBA for 3 days, were infected with bacterial suspension cont...aining 0 or 100 μM AS and than co-cultivated on SIM with the same concentration of AS as infection medium for 3 days. Upon co-cultivation, the explants were cultured on SIM containing 500 mg/l cefotaxime for 8 days, and than subcultured on SIM selection media with 5 mg/l phosphinothricin (PPT) until shoot regeneration. The addition of AS during co-cultivation increased transformation frequency (7.8% vs. 1.4% in its absence). Selected PPT-resistant shoots were continuously maintained and multiplied on MS selection medium containing 2 mg/l PPT and BA or Kin (0.5-1.0 mg/l) alone or in combination with IBA (0.1-0.2 mg/l) displaying vigorous growth and development. Integration of the bar gene in transgenic shoots was verified by chlorophenol red (CR) assay while PCR confirmed the presence of NPK1 gene. Transformed plants provide a promising material for stress tolerance study.
Keywords:
cauliflower / biotechnology / stress tolerance / MAPK / stress responseSource:
1st International conference on Plant Biology and 20th Symposium of the Serbian Plant Physiology Society : Programme and Abstracts, 2011, 64-Publisher:
- Beograd : Serbian Plant Physiology Society
- Beograd : Institute for Biological Research "Siniša Stanković"
Funding / projects:
- Integrating biotechnology approach in breeding vegetable crops for sustainable agricultural systems (RS-31059)
- Biotechnology in vitro - crop, medicinal and endangered plant species (RS-173015)
Collections
Institution/Community
Institut za povrtarstvoTY - CONF AU - Pavlović, Suzana AU - Vinterhalter, Branka AU - Zdravković-Korać, Snežana AU - Adžić, Slađan AU - Đorđević, Mladen AU - Ugrinović, Milan AU - Mitić, Nevena PY - 2011 UR - http://RIVeC.institut-palanka.rs/handle/123456789/577 AB - Cauliflower (Brassica oleracea var. botrytis) is an important vegetable plant grown worldwide that is impacted by biotic and abiotic stresses. Plant biotechnology could be an efficient approach to enhance stress tolerance. Mitogen-activated protein kinase (MAPK) cascades play a key role in plant growth and development as well as biotic and abiotic stress response. Thus, we have established an Agrobaterium tumefacens-mediated protocol for transformation of cauliflower (line KFRM4) using the binary vector pSHX004 with the mitogen-activated protein kinase kinase kinase (MAPKKK) gene from tobacco (NPK1) for stress tolerance and the phosphinothricin acetyltransferase (bar) selectable marker gene for herbicide resistance. The effect of acetosyringone (AS) on transformation efficiency was also investigated. Hypocotyls of 8 day-old seedlings, pre-cultivated on MS shoot induction medium (SIM) supplemented with 1.0 mg/l BA and 0.2 mg/l IBA for 3 days, were infected with bacterial suspension containing 0 or 100 μM AS and than co-cultivated on SIM with the same concentration of AS as infection medium for 3 days. Upon co-cultivation, the explants were cultured on SIM containing 500 mg/l cefotaxime for 8 days, and than subcultured on SIM selection media with 5 mg/l phosphinothricin (PPT) until shoot regeneration. The addition of AS during co-cultivation increased transformation frequency (7.8% vs. 1.4% in its absence). Selected PPT-resistant shoots were continuously maintained and multiplied on MS selection medium containing 2 mg/l PPT and BA or Kin (0.5-1.0 mg/l) alone or in combination with IBA (0.1-0.2 mg/l) displaying vigorous growth and development. Integration of the bar gene in transgenic shoots was verified by chlorophenol red (CR) assay while PCR confirmed the presence of NPK1 gene. Transformed plants provide a promising material for stress tolerance study. PB - Beograd : Serbian Plant Physiology Society PB - Beograd : Institute for Biological Research "Siniša Stanković" C3 - 1st International conference on Plant Biology and 20th Symposium of the Serbian Plant Physiology Society : Programme and Abstracts T1 - Agrobacterium-mediated genetic transformation of cauliflower with the NPK1 gene to enhance biotic and abiotic stress tolerance SP - 64 UR - https://hdl.handle.net/21.15107/rcub_rivec_577 ER -
@conference{ author = "Pavlović, Suzana and Vinterhalter, Branka and Zdravković-Korać, Snežana and Adžić, Slađan and Đorđević, Mladen and Ugrinović, Milan and Mitić, Nevena", year = "2011", abstract = "Cauliflower (Brassica oleracea var. botrytis) is an important vegetable plant grown worldwide that is impacted by biotic and abiotic stresses. Plant biotechnology could be an efficient approach to enhance stress tolerance. Mitogen-activated protein kinase (MAPK) cascades play a key role in plant growth and development as well as biotic and abiotic stress response. Thus, we have established an Agrobaterium tumefacens-mediated protocol for transformation of cauliflower (line KFRM4) using the binary vector pSHX004 with the mitogen-activated protein kinase kinase kinase (MAPKKK) gene from tobacco (NPK1) for stress tolerance and the phosphinothricin acetyltransferase (bar) selectable marker gene for herbicide resistance. The effect of acetosyringone (AS) on transformation efficiency was also investigated. Hypocotyls of 8 day-old seedlings, pre-cultivated on MS shoot induction medium (SIM) supplemented with 1.0 mg/l BA and 0.2 mg/l IBA for 3 days, were infected with bacterial suspension containing 0 or 100 μM AS and than co-cultivated on SIM with the same concentration of AS as infection medium for 3 days. Upon co-cultivation, the explants were cultured on SIM containing 500 mg/l cefotaxime for 8 days, and than subcultured on SIM selection media with 5 mg/l phosphinothricin (PPT) until shoot regeneration. The addition of AS during co-cultivation increased transformation frequency (7.8% vs. 1.4% in its absence). Selected PPT-resistant shoots were continuously maintained and multiplied on MS selection medium containing 2 mg/l PPT and BA or Kin (0.5-1.0 mg/l) alone or in combination with IBA (0.1-0.2 mg/l) displaying vigorous growth and development. Integration of the bar gene in transgenic shoots was verified by chlorophenol red (CR) assay while PCR confirmed the presence of NPK1 gene. Transformed plants provide a promising material for stress tolerance study.", publisher = "Beograd : Serbian Plant Physiology Society, Beograd : Institute for Biological Research "Siniša Stanković"", journal = "1st International conference on Plant Biology and 20th Symposium of the Serbian Plant Physiology Society : Programme and Abstracts", title = "Agrobacterium-mediated genetic transformation of cauliflower with the NPK1 gene to enhance biotic and abiotic stress tolerance", pages = "64", url = "https://hdl.handle.net/21.15107/rcub_rivec_577" }
Pavlović, S., Vinterhalter, B., Zdravković-Korać, S., Adžić, S., Đorđević, M., Ugrinović, M.,& Mitić, N.. (2011). Agrobacterium-mediated genetic transformation of cauliflower with the NPK1 gene to enhance biotic and abiotic stress tolerance. in 1st International conference on Plant Biology and 20th Symposium of the Serbian Plant Physiology Society : Programme and Abstracts Beograd : Serbian Plant Physiology Society., 64. https://hdl.handle.net/21.15107/rcub_rivec_577
Pavlović S, Vinterhalter B, Zdravković-Korać S, Adžić S, Đorđević M, Ugrinović M, Mitić N. Agrobacterium-mediated genetic transformation of cauliflower with the NPK1 gene to enhance biotic and abiotic stress tolerance. in 1st International conference on Plant Biology and 20th Symposium of the Serbian Plant Physiology Society : Programme and Abstracts. 2011;:64. https://hdl.handle.net/21.15107/rcub_rivec_577 .
Pavlović, Suzana, Vinterhalter, Branka, Zdravković-Korać, Snežana, Adžić, Slađan, Đorđević, Mladen, Ugrinović, Milan, Mitić, Nevena, "Agrobacterium-mediated genetic transformation of cauliflower with the NPK1 gene to enhance biotic and abiotic stress tolerance" in 1st International conference on Plant Biology and 20th Symposium of the Serbian Plant Physiology Society : Programme and Abstracts (2011):64, https://hdl.handle.net/21.15107/rcub_rivec_577 .