TY  - CONF
AU  - Simonović, Ana
AU  - Bogdanović, Milica
AU  - Bergau, Nick
AU  - Tissier, Alain
AU  - Todorović, Slađana
PY  - 2018
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/4334
AB  - Tomato (Solanum lycopersicum) is not only one of the most important vegetable crops, but
a widely used model for genetic improvement of dicotyledonous crops. Even though the first
report on tomato transformation using Agrobacterium tumefaciens has been published over 30
years ago, numerous protocols published since then demonstrate that tomato transformation is
neither standardized nor routine. Tomato cultivars and genotypes greatly differ in their susceptibility
to transformation and response to growth regulator and antibiotics combinations used during
regeneration and selection. Hereby we report a successful protocol for “Moneymaker” transformation
with A. tumefaciens GV3101:pSM90 bearing pAGT174 with Kanamycin resistance cassette.
Whole larger leaves (from ~10 cm high plantlets), rather than leaf segments were selected for
inoculation, as the explant size significantly affected regeneration rate. Combination of zeatin (1
mg L-1) and IAA (0.1 mg L-1) was used for preculture and throughout the selection process, with
addition of Timentin (200 mg L-1) and increasing concentrations of Kanamycin (from 20 to 50 mg
L 1) during the selection and regeneration. The regeneration on Kan occurred almost exclusively
in transformed explants and proceeded via organogenesis with the callusing stage. Using Timentin
instead of commonly used Cefotaxime to kill the bacteria proved beneficial for Moneymaker
regeneration. In this system the regeneration frequency was 7%. Plantlets that survived Kan selection
were PCR-tested, and 89.6% of them were transformed. The protocol is convenient and robust
in terms of very low false-positive rate (10.3%).
PB  - Belgrade: Serbian Plant Physiology Society
C3  - 3rd International Conference on Plant Biology (22nd SPPS Meeting), Belgrade
T1  - Transformation of tomato cultivar Moneymaker with Agrobacterium tumefaciens
SP  - 21
UR  - https://hdl.handle.net/21.15107/rcub_ibiss_4334
ER  - 

@conference{
author = "Simonović, Ana and Bogdanović, Milica and Bergau, Nick and Tissier, Alain and Todorović, Slađana",
year = "2018",
abstract = "Tomato (Solanum lycopersicum) is not only one of the most important vegetable crops, but
a widely used model for genetic improvement of dicotyledonous crops. Even though the first
report on tomato transformation using Agrobacterium tumefaciens has been published over 30
years ago, numerous protocols published since then demonstrate that tomato transformation is
neither standardized nor routine. Tomato cultivars and genotypes greatly differ in their susceptibility
to transformation and response to growth regulator and antibiotics combinations used during
regeneration and selection. Hereby we report a successful protocol for “Moneymaker” transformation
with A. tumefaciens GV3101:pSM90 bearing pAGT174 with Kanamycin resistance cassette.
Whole larger leaves (from ~10 cm high plantlets), rather than leaf segments were selected for
inoculation, as the explant size significantly affected regeneration rate. Combination of zeatin (1
mg L-1) and IAA (0.1 mg L-1) was used for preculture and throughout the selection process, with
addition of Timentin (200 mg L-1) and increasing concentrations of Kanamycin (from 20 to 50 mg
L 1) during the selection and regeneration. The regeneration on Kan occurred almost exclusively
in transformed explants and proceeded via organogenesis with the callusing stage. Using Timentin
instead of commonly used Cefotaxime to kill the bacteria proved beneficial for Moneymaker
regeneration. In this system the regeneration frequency was 7%. Plantlets that survived Kan selection
were PCR-tested, and 89.6% of them were transformed. The protocol is convenient and robust
in terms of very low false-positive rate (10.3%).",
publisher = "Belgrade: Serbian Plant Physiology Society",
journal = "3rd International Conference on Plant Biology (22nd SPPS Meeting), Belgrade",
title = "Transformation of tomato cultivar Moneymaker with Agrobacterium tumefaciens",
pages = "21",
url = "https://hdl.handle.net/21.15107/rcub_ibiss_4334"
}

Simonović, A., Bogdanović, M., Bergau, N., Tissier, A.,& Todorović, S.. (2018). Transformation of tomato cultivar Moneymaker with Agrobacterium tumefaciens. in 3rd International Conference on Plant Biology (22nd SPPS Meeting), Belgrade
Belgrade: Serbian Plant Physiology Society., 21.
https://hdl.handle.net/21.15107/rcub_ibiss_4334

Simonović A, Bogdanović M, Bergau N, Tissier A, Todorović S. Transformation of tomato cultivar Moneymaker with Agrobacterium tumefaciens. in 3rd International Conference on Plant Biology (22nd SPPS Meeting), Belgrade. 2018;:21.
https://hdl.handle.net/21.15107/rcub_ibiss_4334 .

Simonović, Ana, Bogdanović, Milica, Bergau, Nick, Tissier, Alain, Todorović, Slađana, "Transformation of tomato cultivar Moneymaker with Agrobacterium tumefaciens" in 3rd International Conference on Plant Biology (22nd SPPS Meeting), Belgrade (2018):21,
https://hdl.handle.net/21.15107/rcub_ibiss_4334 .

TY  - JOUR
AU  - Brückner, Kathleen
AU  - Matekalo, Dragana
AU  - Manzano, David
AU  - Papaefthimiou, Dimitra
AU  - Pateraki, Irini
AU  - Scheler, Ulschan
AU  - Ferrer, Albert
AU  - de Vos, Ric C H
AU  - Kanellis, Angelos
AU  - Tissier, Alain
PY  - 2014
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/4287
AB  - Rosemary (Rosmarinus officinalis) produces the phenolic diterpenes carnosic acid and carnosol, which, in addition to their general antioxidant activities, have recently been suggested as potential ingredients for the prevention and treatment of neurodegenerative diseases. Little is known about the biosynthesis of these diterpenes. Here we show that the biosynthesis of phenolic diterpenes in rosemary predominantly takes place in the glandular trichomes of young leaves, and used this feature to identify the first committed steps. Thus, a copalyl diphosphate synthase (RoCPS1) and two kaurene synthase-like (RoKSL1 and RoKSL2) encoding genes were identified and characterized. Expression in yeast (Saccharomyces cerevisiae) and Nicotiana benthamiana demonstrate that RoCPS1 converts geranylgeranyl diphosphate (GGDP) to copalyl diphosphate (CDP) of normal stereochemistry and that both RoKSL1 and RoKSL2 use normal CDP to produce an abietane diterpene. Comparison to the already characterized diterpene synthase from Salvia miltiorrhiza (SmKSL) demonstrates that the product of RoKSL1 and RoKSL2 is miltiradiene. Expression analysis supports a major contributing role for RoKSL2. Like SmKSL and the sclareol synthase from Salvia sclarea, RoKSL1/2 are diterpene synthases of the TPS-e group which have lost the internal gamma-domain. Furthermore, phylogenetic analysis indicates that RoKSL1 and RoKSL2 belong to a distinct group of KSL enzymes involved in specialized metabolism which most likely emerged before the dicot-monocot split.
PB  - Elsevier
T2  - Phytochemistry
T1  - Characterization of two genes for the biosynthesis of abietane-type diterpenes in rosemary (Rosmarinus officinalis) glandular trichomes
VL  - 101
DO  - 10.1016/j.phytochem.2014.01.021
SP  - 52
EP  - 64
ER  - 

@article{
author = "Brückner, Kathleen and Matekalo, Dragana and Manzano, David and Papaefthimiou, Dimitra and Pateraki, Irini and Scheler, Ulschan and Ferrer, Albert and de Vos, Ric C H and Kanellis, Angelos and Tissier, Alain",
year = "2014",
abstract = "Rosemary (Rosmarinus officinalis) produces the phenolic diterpenes carnosic acid and carnosol, which, in addition to their general antioxidant activities, have recently been suggested as potential ingredients for the prevention and treatment of neurodegenerative diseases. Little is known about the biosynthesis of these diterpenes. Here we show that the biosynthesis of phenolic diterpenes in rosemary predominantly takes place in the glandular trichomes of young leaves, and used this feature to identify the first committed steps. Thus, a copalyl diphosphate synthase (RoCPS1) and two kaurene synthase-like (RoKSL1 and RoKSL2) encoding genes were identified and characterized. Expression in yeast (Saccharomyces cerevisiae) and Nicotiana benthamiana demonstrate that RoCPS1 converts geranylgeranyl diphosphate (GGDP) to copalyl diphosphate (CDP) of normal stereochemistry and that both RoKSL1 and RoKSL2 use normal CDP to produce an abietane diterpene. Comparison to the already characterized diterpene synthase from Salvia miltiorrhiza (SmKSL) demonstrates that the product of RoKSL1 and RoKSL2 is miltiradiene. Expression analysis supports a major contributing role for RoKSL2. Like SmKSL and the sclareol synthase from Salvia sclarea, RoKSL1/2 are diterpene synthases of the TPS-e group which have lost the internal gamma-domain. Furthermore, phylogenetic analysis indicates that RoKSL1 and RoKSL2 belong to a distinct group of KSL enzymes involved in specialized metabolism which most likely emerged before the dicot-monocot split.",
publisher = "Elsevier",
journal = "Phytochemistry",
title = "Characterization of two genes for the biosynthesis of abietane-type diterpenes in rosemary (Rosmarinus officinalis) glandular trichomes",
volume = "101",
doi = "10.1016/j.phytochem.2014.01.021",
pages = "52-64"
}

Brückner, K., Matekalo, D., Manzano, D., Papaefthimiou, D., Pateraki, I., Scheler, U., Ferrer, A., de Vos, R. C. H., Kanellis, A.,& Tissier, A.. (2014). Characterization of two genes for the biosynthesis of abietane-type diterpenes in rosemary (Rosmarinus officinalis) glandular trichomes. in Phytochemistry
Elsevier., 101, 52-64.
https://doi.org/10.1016/j.phytochem.2014.01.021

Brückner K, Matekalo D, Manzano D, Papaefthimiou D, Pateraki I, Scheler U, Ferrer A, de Vos RCH, Kanellis A, Tissier A. Characterization of two genes for the biosynthesis of abietane-type diterpenes in rosemary (Rosmarinus officinalis) glandular trichomes. in Phytochemistry. 2014;101:52-64.
doi:10.1016/j.phytochem.2014.01.021 .

Brückner, Kathleen, Matekalo, Dragana, Manzano, David, Papaefthimiou, Dimitra, Pateraki, Irini, Scheler, Ulschan, Ferrer, Albert, de Vos, Ric C H, Kanellis, Angelos, Tissier, Alain, "Characterization of two genes for the biosynthesis of abietane-type diterpenes in rosemary (Rosmarinus officinalis) glandular trichomes" in Phytochemistry, 101 (2014):52-64,
https://doi.org/10.1016/j.phytochem.2014.01.021 . .

TY  - CONF
AU  - Brückner, Kathleen
AU  - Matekalo, Dragana
AU  - Manzano, David
AU  - Papaefthimiou, Dimitra
AU  - Pateraki, Irini
AU  - Boronat, Albert
AU  - Datema, Erwin
AU  - de Vos, Ric C H
AU  - Ferrer, Albert
AU  - Kanellis, Angelos
AU  - Tissier, Alain
PY  - 2013
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/4288
AB  - Plant secondary metabolites contribute substantially to the human pharmacopeae. To better exploit the medicinal plants that produce these compounds, it is necessary to understand how these compounds are synthesized within the plant. Identification of the biosynthetic pathway genes will allow both the breeding of plant varieties with improved productivity and/or metabolic profile, and the development of alternative production methods using biotechnology.
The phenolic diterpenes (PDs) carnosic acid (CA) and carnosol have high antioxidant activities and display potential for the treatment of neurodegenerative disorders. CA and carnosol are found in Rosmarinus and Salvia sp. which are known for their health promoting properties since ages.
As part of the EU-financed TERPMED project, PD biosynthesis is being investigated. To elucidate the pathway, the localization of the biosynthesis of PDs was determined. Glandular trichomes appear to contribute significantly to the biosynthesis of PDs and the transcriptome of glandular trichomes was thus determined by next generation sequencing of cDNA.
A set of gene candidates was selected according to alignment searches and analysis of trichome-specific gene expression profile. The search for terpene synthases yielded two types of candidates for the first steps of the pathway: a copalyl diphosphate synthase (CPS) and kaurene synthase-like (KSL) encoding genes. Genes encoding oxidases of cytochrome P450 clade were the most promising candidates for the downstream steps of the PDs pathway.
Finally, the first steps of the biosynthesis of PDs are reconstituted and further investigations on the role of P450 candidates in downstream steps are being carried out using N. benthamiana transient expression and heterologous expression in yeast.
PB  - Thessaloniki: Department of Pharmaceutical Science, Aristotle University of Thessaloniki
C3  - 11th International meeting Biosynthesis, Function and Biotechnology of Isoprenoids in Terrestrial and Marine Organisms, Book of Abstracts
T1  - Investigating phenolic diterpenes in rosemary (Rosmarinus officinalis) and sage (Salvia fruticosa)
SP  - 59
UR  - https://hdl.handle.net/21.15107/rcub_ibiss_4288
ER  - 

@conference{
author = "Brückner, Kathleen and Matekalo, Dragana and Manzano, David and Papaefthimiou, Dimitra and Pateraki, Irini and Boronat, Albert and Datema, Erwin and de Vos, Ric C H and Ferrer, Albert and Kanellis, Angelos and Tissier, Alain",
year = "2013",
abstract = "Plant secondary metabolites contribute substantially to the human pharmacopeae. To better exploit the medicinal plants that produce these compounds, it is necessary to understand how these compounds are synthesized within the plant. Identification of the biosynthetic pathway genes will allow both the breeding of plant varieties with improved productivity and/or metabolic profile, and the development of alternative production methods using biotechnology.
The phenolic diterpenes (PDs) carnosic acid (CA) and carnosol have high antioxidant activities and display potential for the treatment of neurodegenerative disorders. CA and carnosol are found in Rosmarinus and Salvia sp. which are known for their health promoting properties since ages.
As part of the EU-financed TERPMED project, PD biosynthesis is being investigated. To elucidate the pathway, the localization of the biosynthesis of PDs was determined. Glandular trichomes appear to contribute significantly to the biosynthesis of PDs and the transcriptome of glandular trichomes was thus determined by next generation sequencing of cDNA.
A set of gene candidates was selected according to alignment searches and analysis of trichome-specific gene expression profile. The search for terpene synthases yielded two types of candidates for the first steps of the pathway: a copalyl diphosphate synthase (CPS) and kaurene synthase-like (KSL) encoding genes. Genes encoding oxidases of cytochrome P450 clade were the most promising candidates for the downstream steps of the PDs pathway.
Finally, the first steps of the biosynthesis of PDs are reconstituted and further investigations on the role of P450 candidates in downstream steps are being carried out using N. benthamiana transient expression and heterologous expression in yeast.",
publisher = "Thessaloniki: Department of Pharmaceutical Science, Aristotle University of Thessaloniki",
journal = "11th International meeting Biosynthesis, Function and Biotechnology of Isoprenoids in Terrestrial and Marine Organisms, Book of Abstracts",
title = "Investigating phenolic diterpenes in rosemary (Rosmarinus officinalis) and sage (Salvia fruticosa)",
pages = "59",
url = "https://hdl.handle.net/21.15107/rcub_ibiss_4288"
}

Brückner, K., Matekalo, D., Manzano, D., Papaefthimiou, D., Pateraki, I., Boronat, A., Datema, E., de Vos, R. C. H., Ferrer, A., Kanellis, A.,& Tissier, A.. (2013). Investigating phenolic diterpenes in rosemary (Rosmarinus officinalis) and sage (Salvia fruticosa). in 11th International meeting Biosynthesis, Function and Biotechnology of Isoprenoids in Terrestrial and Marine Organisms, Book of Abstracts
Thessaloniki: Department of Pharmaceutical Science, Aristotle University of Thessaloniki., 59.
https://hdl.handle.net/21.15107/rcub_ibiss_4288

Brückner K, Matekalo D, Manzano D, Papaefthimiou D, Pateraki I, Boronat A, Datema E, de Vos RCH, Ferrer A, Kanellis A, Tissier A. Investigating phenolic diterpenes in rosemary (Rosmarinus officinalis) and sage (Salvia fruticosa). in 11th International meeting Biosynthesis, Function and Biotechnology of Isoprenoids in Terrestrial and Marine Organisms, Book of Abstracts. 2013;:59.
https://hdl.handle.net/21.15107/rcub_ibiss_4288 .

Brückner, Kathleen, Matekalo, Dragana, Manzano, David, Papaefthimiou, Dimitra, Pateraki, Irini, Boronat, Albert, Datema, Erwin, de Vos, Ric C H, Ferrer, Albert, Kanellis, Angelos, Tissier, Alain, "Investigating phenolic diterpenes in rosemary (Rosmarinus officinalis) and sage (Salvia fruticosa)" in 11th International meeting Biosynthesis, Function and Biotechnology of Isoprenoids in Terrestrial and Marine Organisms, Book of Abstracts (2013):59,
https://hdl.handle.net/21.15107/rcub_ibiss_4288 .

TY  - JOUR
AU  - Bogdanović, Milica
AU  - Dragičević, Milan B.
AU  - Tanić, Nikola T
AU  - Todorović, Slađana
AU  - Mišić, Danijela
AU  - Živković, Suzana
AU  - Tissier, Alain
AU  - Simonović, Ana
PY  - 2013
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/1045
AB  - Ribosomal 18S RNA is widely used as a housekeeping gene in expression studies, including end-point PCR, Northern analysis, and real-time experiments. However, there are two disadvantages and two points of error introduction in using 18S rRNA as a reference gene. First, 18S has no poly(A) tail, so it is commonly reverse transcribed with specific primers or random hexamers, independently from poly(dT)-primed transcripts. Secondly, due to its abundance, the 18S cDNA must be extensively diluted to be comparable to the tested genes. In this study, 18S rRNA from five taxonomically diverse plant species, including Physcomitrella patens, Adiantum capillus-veneris, Centaurium erythraea, Arabidopsis thaliana, and Zea mays, was successfully reverse transcribed (RT) using poly(dT)(18). As all other homopolymers, including poly(dA)(18), poly(dC)(18), and poly(dG)(18), could serve as RT primers, it was concluded that homopolymers anneal by mispriming at the sites of complementary homopolymeric runs or segments rich in complementary base. Poly(dC)(18) was the most efficient as RT primer, and the only one which interfered with subsequent PCR, giving species-specific pattern of products. Poly(dT)-primed RT reactions were less efficient in comparison to specific primer or random hexamer-primed reactions. Homopolymeric priming of 18S in RT reactions is general in terms of RNA origin and the method of RNA isolation and is possibly applicable to other tailless housekeeping genes.
T2  - Plant Molecular Biology Reporter
T1  - Reverse Transcription of 18S rRNA with Poly(dT)(18) and Other Homopolymers
IS  - 1
VL  - 31
SP  - 1
EP  - 63
UR  - https://hdl.handle.net/21.15107/rcub_ibiss_1045
ER  - 

@article{
author = "Bogdanović, Milica and Dragičević, Milan B. and Tanić, Nikola T and Todorović, Slađana and Mišić, Danijela and Živković, Suzana and Tissier, Alain and Simonović, Ana",
year = "2013",
abstract = "Ribosomal 18S RNA is widely used as a housekeeping gene in expression studies, including end-point PCR, Northern analysis, and real-time experiments. However, there are two disadvantages and two points of error introduction in using 18S rRNA as a reference gene. First, 18S has no poly(A) tail, so it is commonly reverse transcribed with specific primers or random hexamers, independently from poly(dT)-primed transcripts. Secondly, due to its abundance, the 18S cDNA must be extensively diluted to be comparable to the tested genes. In this study, 18S rRNA from five taxonomically diverse plant species, including Physcomitrella patens, Adiantum capillus-veneris, Centaurium erythraea, Arabidopsis thaliana, and Zea mays, was successfully reverse transcribed (RT) using poly(dT)(18). As all other homopolymers, including poly(dA)(18), poly(dC)(18), and poly(dG)(18), could serve as RT primers, it was concluded that homopolymers anneal by mispriming at the sites of complementary homopolymeric runs or segments rich in complementary base. Poly(dC)(18) was the most efficient as RT primer, and the only one which interfered with subsequent PCR, giving species-specific pattern of products. Poly(dT)-primed RT reactions were less efficient in comparison to specific primer or random hexamer-primed reactions. Homopolymeric priming of 18S in RT reactions is general in terms of RNA origin and the method of RNA isolation and is possibly applicable to other tailless housekeeping genes.",
journal = "Plant Molecular Biology Reporter",
title = "Reverse Transcription of 18S rRNA with Poly(dT)(18) and Other Homopolymers",
number = "1",
volume = "31",
pages = "1-63",
url = "https://hdl.handle.net/21.15107/rcub_ibiss_1045"
}

Bogdanović, M., Dragičević, M. B., Tanić, N. T., Todorović, S., Mišić, D., Živković, S., Tissier, A.,& Simonović, A.. (2013). Reverse Transcription of 18S rRNA with Poly(dT)(18) and Other Homopolymers. in Plant Molecular Biology Reporter, 31(1), 1-63.
https://hdl.handle.net/21.15107/rcub_ibiss_1045

Bogdanović M, Dragičević MB, Tanić NT, Todorović S, Mišić D, Živković S, Tissier A, Simonović A. Reverse Transcription of 18S rRNA with Poly(dT)(18) and Other Homopolymers. in Plant Molecular Biology Reporter. 2013;31(1):1-63.
https://hdl.handle.net/21.15107/rcub_ibiss_1045 .

Bogdanović, Milica, Dragičević, Milan B., Tanić, Nikola T, Todorović, Slađana, Mišić, Danijela, Živković, Suzana, Tissier, Alain, Simonović, Ana, "Reverse Transcription of 18S rRNA with Poly(dT)(18) and Other Homopolymers" in Plant Molecular Biology Reporter, 31, no. 1 (2013):1-63,
https://hdl.handle.net/21.15107/rcub_ibiss_1045 .