TY  - JOUR
AU  - Pavlović, Suzana
AU  - Savić, Jelena
AU  - Milojević, Jelena
AU  - Vinterhalter, Branka
AU  - Girek, Zdenka
AU  - Adžić, Slađan
AU  - Zečević, Bogoljub
AU  - Banjac, Nevena
PY  - 2020
UR  - http://RIVeC.institut-palanka.rs/handle/123456789/215
AB  - Key message The results underlined that constitutively expressingNPK1can significantly contribute to enhanced salt stress tolerance in cauliflower, suggesting that this could be a promising basis for the creation of new stress tolerance cruciferous vegetable lines. Cauliflower is exposed to various biotic and abiotic stresses, including increased salinity due to the intensive irrigation of crops. Mitogen-activated protein kinase (MAPK) cascades are universal signal transduction modules that play important roles in regulating innate immune responses in plants. Based on involvement of tobacco MAP kinase kinase kinase (NPK1) in stress response, the effect of the expression ofNPK1transgene to NaCl salt stress tolerance in cauliflower KFRM4 lines was studied. TheAgrobacterium tumefaciens-mediated transformation protocol, using EHA101(pSHX004) vector harbouring theNPK1and phosphinothricinN-acetyltransferase (bar) genes, the cyclic somatic embryogenesis regeneration pathway, the application of acetosyringone (AS) during co-cultivation and a delayed phosphinothricine (PPT) selection procedure provided sufficient transformation efficiency of 7.33% without escapes. PCR analysis indicated the integration of bothNPK1andbartransgenes in regenerated cauliflower lines. Transgenic cauliflower lines, exposed to NaCl stress in vitro, showed higher growth rates, greater ability to retain chlorophyll and carotenoids, and increased osmotic regulation capacity compared with non-transformed control plants. The tolerance level of transformed lines correlated with the level ofNPK1gene expression estimated by RT-qPCR, and the L2 line with the highestNPK1expression displayed the greatest tolerance to NaCl stress. None of the obtained cauliflower transformed lines grown in greenhouses showed any morphological or yield differences compared with non-transformed plants. Furthermore, the expression of thebargene facilitated the tolerance of transformed lines to the total herbicide PPT, applied at concentrations 2-3 times higher than those routinely used for weed control in the crop field.
PB  - Springer, Dordrecht
T2  - Plant Cell Tissue and Organ Culture
T1  - Introduction of the Nicotiana protein kinase (NPK1) gene by combiningAgrobacterium-mediated transformation and recurrent somatic embryogenesis to enhance salt tolerance in cauliflower
EP  - 651
IS  - 3
SP  - 635
VL  - 143
DO  - 10.1007/s11240-020-01948-6
ER  - 

@article{
author = "Pavlović, Suzana and Savić, Jelena and Milojević, Jelena and Vinterhalter, Branka and Girek, Zdenka and Adžić, Slađan and Zečević, Bogoljub and Banjac, Nevena",
year = "2020",
abstract = "Key message The results underlined that constitutively expressingNPK1can significantly contribute to enhanced salt stress tolerance in cauliflower, suggesting that this could be a promising basis for the creation of new stress tolerance cruciferous vegetable lines. Cauliflower is exposed to various biotic and abiotic stresses, including increased salinity due to the intensive irrigation of crops. Mitogen-activated protein kinase (MAPK) cascades are universal signal transduction modules that play important roles in regulating innate immune responses in plants. Based on involvement of tobacco MAP kinase kinase kinase (NPK1) in stress response, the effect of the expression ofNPK1transgene to NaCl salt stress tolerance in cauliflower KFRM4 lines was studied. TheAgrobacterium tumefaciens-mediated transformation protocol, using EHA101(pSHX004) vector harbouring theNPK1and phosphinothricinN-acetyltransferase (bar) genes, the cyclic somatic embryogenesis regeneration pathway, the application of acetosyringone (AS) during co-cultivation and a delayed phosphinothricine (PPT) selection procedure provided sufficient transformation efficiency of 7.33% without escapes. PCR analysis indicated the integration of bothNPK1andbartransgenes in regenerated cauliflower lines. Transgenic cauliflower lines, exposed to NaCl stress in vitro, showed higher growth rates, greater ability to retain chlorophyll and carotenoids, and increased osmotic regulation capacity compared with non-transformed control plants. The tolerance level of transformed lines correlated with the level ofNPK1gene expression estimated by RT-qPCR, and the L2 line with the highestNPK1expression displayed the greatest tolerance to NaCl stress. None of the obtained cauliflower transformed lines grown in greenhouses showed any morphological or yield differences compared with non-transformed plants. Furthermore, the expression of thebargene facilitated the tolerance of transformed lines to the total herbicide PPT, applied at concentrations 2-3 times higher than those routinely used for weed control in the crop field.",
publisher = "Springer, Dordrecht",
journal = "Plant Cell Tissue and Organ Culture",
title = "Introduction of the Nicotiana protein kinase (NPK1) gene by combiningAgrobacterium-mediated transformation and recurrent somatic embryogenesis to enhance salt tolerance in cauliflower",
pages = "651-635",
number = "3",
volume = "143",
doi = "10.1007/s11240-020-01948-6"
}

Pavlović, S., Savić, J., Milojević, J., Vinterhalter, B., Girek, Z., Adžić, S., Zečević, B.,& Banjac, N.. (2020). Introduction of the Nicotiana protein kinase (NPK1) gene by combiningAgrobacterium-mediated transformation and recurrent somatic embryogenesis to enhance salt tolerance in cauliflower. in Plant Cell Tissue and Organ Culture
Springer, Dordrecht., 143(3), 635-651.
https://doi.org/10.1007/s11240-020-01948-6

Pavlović S, Savić J, Milojević J, Vinterhalter B, Girek Z, Adžić S, Zečević B, Banjac N. Introduction of the Nicotiana protein kinase (NPK1) gene by combiningAgrobacterium-mediated transformation and recurrent somatic embryogenesis to enhance salt tolerance in cauliflower. in Plant Cell Tissue and Organ Culture. 2020;143(3):635-651.
doi:10.1007/s11240-020-01948-6 .

Pavlović, Suzana, Savić, Jelena, Milojević, Jelena, Vinterhalter, Branka, Girek, Zdenka, Adžić, Slađan, Zečević, Bogoljub, Banjac, Nevena, "Introduction of the Nicotiana protein kinase (NPK1) gene by combiningAgrobacterium-mediated transformation and recurrent somatic embryogenesis to enhance salt tolerance in cauliflower" in Plant Cell Tissue and Organ Culture, 143, no. 3 (2020):635-651,
https://doi.org/10.1007/s11240-020-01948-6 . .

TY  - JOUR
AU  - Pavlović, Suzana
AU  - Vinterhalter, Branka
AU  - Zdravković-Korać, Snežana
AU  - Vinterhalter, Dragan
AU  - Zdravković, Jasmina
AU  - Cvikić, Dejan
AU  - Mitić, Nevena
PY  - 2013
UR  - http://RIVeC.institut-palanka.rs/handle/123456789/143
AB  - A simple and rapid protocol was established for repetitive somatic embryogenesis and subsequent plant regeneration in two important Brassica oleracea varieties, cabbage and cauliflower. Direct regeneration of somatic embryos (SEs) was achieved from immature zygotic embryos cultured on B5 plant growth regulator (PGR)-free (B5-0) induction medium and on B5 medium supplemented with 1 mg l(-1) 2,4-dichlorophenoxyacetic acid (2,4-D) (B5-D). Zygotic embryos of both cabbage and cauliflower at the cotyledonary (C) stage (1.8 mm long) incubated on B5-0 medium displayed the highest embryo-forming capacities (EFCs) of 11.84 and 11.95, respectively. Secondary somatic embryos (SSEs) appeared on the cabbage and cauliflower's primary embryos at a high frequency (83.3 and 87.5 %, respectively), and this process continued in a repetitive way on PGR-free Murashige and Skoog (MS-0) medium. The embryogenic potential of the cultures with a gradual diminution was maintained for 10 months (ten cycles). A total of 20 % of the mature SSEs from cabbage and 55 % from cauliflower spontaneously regenerated plantlets on MS-0 medium. The addition of 1 mg l(-1) 6-benzyladenine (BA) or 6-furfurylaminopurine (Kin) in the regeneration medium significantly improved somatic embryo conversion into plantlets by up to 56 % in cabbage and 79 % in cauliflower. Regenerated plants acclimated successfully to ex vitro conditions and displayed morphological and reproductive characteristics similar to seed-derived plants. Effective recurrent somatic embryogenesis may be an appropriate practical solution for clonal propagation and genetic modifications of cabbage and cauliflower.
PB  - Springer, Dordrecht
T2  - Plant Cell Tissue and Organ Culture
T1  - Recurrent somatic embryogenesis and plant regeneration from immature zygotic embryos of cabbage (Brassica oleracea var. capitata) and cauliflower (Brassica oleracea var. botrytis)
EP  - 406
IS  - 3
SP  - 397
VL  - 113
DO  - 10.1007/s11240-012-0279-6
ER  - 

@article{
author = "Pavlović, Suzana and Vinterhalter, Branka and Zdravković-Korać, Snežana and Vinterhalter, Dragan and Zdravković, Jasmina and Cvikić, Dejan and Mitić, Nevena",
year = "2013",
abstract = "A simple and rapid protocol was established for repetitive somatic embryogenesis and subsequent plant regeneration in two important Brassica oleracea varieties, cabbage and cauliflower. Direct regeneration of somatic embryos (SEs) was achieved from immature zygotic embryos cultured on B5 plant growth regulator (PGR)-free (B5-0) induction medium and on B5 medium supplemented with 1 mg l(-1) 2,4-dichlorophenoxyacetic acid (2,4-D) (B5-D). Zygotic embryos of both cabbage and cauliflower at the cotyledonary (C) stage (1.8 mm long) incubated on B5-0 medium displayed the highest embryo-forming capacities (EFCs) of 11.84 and 11.95, respectively. Secondary somatic embryos (SSEs) appeared on the cabbage and cauliflower's primary embryos at a high frequency (83.3 and 87.5 %, respectively), and this process continued in a repetitive way on PGR-free Murashige and Skoog (MS-0) medium. The embryogenic potential of the cultures with a gradual diminution was maintained for 10 months (ten cycles). A total of 20 % of the mature SSEs from cabbage and 55 % from cauliflower spontaneously regenerated plantlets on MS-0 medium. The addition of 1 mg l(-1) 6-benzyladenine (BA) or 6-furfurylaminopurine (Kin) in the regeneration medium significantly improved somatic embryo conversion into plantlets by up to 56 % in cabbage and 79 % in cauliflower. Regenerated plants acclimated successfully to ex vitro conditions and displayed morphological and reproductive characteristics similar to seed-derived plants. Effective recurrent somatic embryogenesis may be an appropriate practical solution for clonal propagation and genetic modifications of cabbage and cauliflower.",
publisher = "Springer, Dordrecht",
journal = "Plant Cell Tissue and Organ Culture",
title = "Recurrent somatic embryogenesis and plant regeneration from immature zygotic embryos of cabbage (Brassica oleracea var. capitata) and cauliflower (Brassica oleracea var. botrytis)",
pages = "406-397",
number = "3",
volume = "113",
doi = "10.1007/s11240-012-0279-6"
}

Pavlović, S., Vinterhalter, B., Zdravković-Korać, S., Vinterhalter, D., Zdravković, J., Cvikić, D.,& Mitić, N.. (2013). Recurrent somatic embryogenesis and plant regeneration from immature zygotic embryos of cabbage (Brassica oleracea var. capitata) and cauliflower (Brassica oleracea var. botrytis). in Plant Cell Tissue and Organ Culture
Springer, Dordrecht., 113(3), 397-406.
https://doi.org/10.1007/s11240-012-0279-6

Pavlović S, Vinterhalter B, Zdravković-Korać S, Vinterhalter D, Zdravković J, Cvikić D, Mitić N. Recurrent somatic embryogenesis and plant regeneration from immature zygotic embryos of cabbage (Brassica oleracea var. capitata) and cauliflower (Brassica oleracea var. botrytis). in Plant Cell Tissue and Organ Culture. 2013;113(3):397-406.
doi:10.1007/s11240-012-0279-6 .

Pavlović, Suzana, Vinterhalter, Branka, Zdravković-Korać, Snežana, Vinterhalter, Dragan, Zdravković, Jasmina, Cvikić, Dejan, Mitić, Nevena, "Recurrent somatic embryogenesis and plant regeneration from immature zygotic embryos of cabbage (Brassica oleracea var. capitata) and cauliflower (Brassica oleracea var. botrytis)" in Plant Cell Tissue and Organ Culture, 113, no. 3 (2013):397-406,
https://doi.org/10.1007/s11240-012-0279-6 . .

TY  - JOUR
AU  - Milojević, Jelena
AU  - Tubić, Ljiljana
AU  - Pavlović, Suzana
AU  - Mitić, Nevena
AU  - Calić, Dušica
AU  - Vinterhalter, Branka
AU  - Zdravković-Korać, Snežana
PY  - 2012
UR  - http://RIVeC.institut-palanka.rs/handle/123456789/123
AB  - As has been previously shown, the genotype at the individual plant level has a strong effect on the regeneration capacity of spinach, and this effect quite often masks the impact of other factors, including the light environment. Therefore, apical sections of the lateral roots of the same seedling (line) were exposed in parallel to both long day (LD, 16-h photoperiod) and short day (SD, 8-h photoperiod) conditions to assess the effect of photoperiod on somatic embryo (SE) regeneration. The root sections in most lines responded with significantly higher frequencies of regeneration and mean SE numbers with LD compared to roots of the same origin with SD. while 7 of 40 lines were only able to regenerate when exposed to LD. Moreover, in all lines cultured with SD, regeneration was delayed by approximately 4 weeks. Obtained SEs were further grown and multiplied, providing material for the evaluation of the effect of light intensity on regeneration capacity. Maximal regeneration capacity was recorded at 100 mu mol m(-2) s(-1) in all lines tested. The present research confirmed the previously observed strong effect of genotype on regeneration capacity and undoubtedly showed a significantly better response in explants exposed to LD than to SD. These results could contribute to better understanding the process of SE induction in spinach and further improve the efficiency of this process.
PB  - Elsevier Science Bv, Amsterdam
T2  - Scientia Horticulturae
T1  - Long days promote somatic embryogenesis in spinach
EP  - 37
SP  - 32
VL  - 142
DO  - 10.1016/j.scienta.2012.04.020
ER  - 

@article{
author = "Milojević, Jelena and Tubić, Ljiljana and Pavlović, Suzana and Mitić, Nevena and Calić, Dušica and Vinterhalter, Branka and Zdravković-Korać, Snežana",
year = "2012",
abstract = "As has been previously shown, the genotype at the individual plant level has a strong effect on the regeneration capacity of spinach, and this effect quite often masks the impact of other factors, including the light environment. Therefore, apical sections of the lateral roots of the same seedling (line) were exposed in parallel to both long day (LD, 16-h photoperiod) and short day (SD, 8-h photoperiod) conditions to assess the effect of photoperiod on somatic embryo (SE) regeneration. The root sections in most lines responded with significantly higher frequencies of regeneration and mean SE numbers with LD compared to roots of the same origin with SD. while 7 of 40 lines were only able to regenerate when exposed to LD. Moreover, in all lines cultured with SD, regeneration was delayed by approximately 4 weeks. Obtained SEs were further grown and multiplied, providing material for the evaluation of the effect of light intensity on regeneration capacity. Maximal regeneration capacity was recorded at 100 mu mol m(-2) s(-1) in all lines tested. The present research confirmed the previously observed strong effect of genotype on regeneration capacity and undoubtedly showed a significantly better response in explants exposed to LD than to SD. These results could contribute to better understanding the process of SE induction in spinach and further improve the efficiency of this process.",
publisher = "Elsevier Science Bv, Amsterdam",
journal = "Scientia Horticulturae",
title = "Long days promote somatic embryogenesis in spinach",
pages = "37-32",
volume = "142",
doi = "10.1016/j.scienta.2012.04.020"
}

Milojević, J., Tubić, L., Pavlović, S., Mitić, N., Calić, D., Vinterhalter, B.,& Zdravković-Korać, S.. (2012). Long days promote somatic embryogenesis in spinach. in Scientia Horticulturae
Elsevier Science Bv, Amsterdam., 142, 32-37.
https://doi.org/10.1016/j.scienta.2012.04.020

Milojević J, Tubić L, Pavlović S, Mitić N, Calić D, Vinterhalter B, Zdravković-Korać S. Long days promote somatic embryogenesis in spinach. in Scientia Horticulturae. 2012;142:32-37.
doi:10.1016/j.scienta.2012.04.020 .

Milojević, Jelena, Tubić, Ljiljana, Pavlović, Suzana, Mitić, Nevena, Calić, Dušica, Vinterhalter, Branka, Zdravković-Korać, Snežana, "Long days promote somatic embryogenesis in spinach" in Scientia Horticulturae, 142 (2012):32-37,
https://doi.org/10.1016/j.scienta.2012.04.020 . .

TY  - 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 .