Immature Zygotic Embryo Cultures of Arabidopsis. A Model System for Molecular Studies on Morphogenic Pathways Induced In Vitro

Kraut, Małgorzata; Wójcikowska, Barbara; Ledwoń, Agnieszka; Gaj, Małgorzata D.

doi:10.2478/v10182-011-0028-x

Cited by 7 publications

(13 citation statements)

References 50 publications

(43 reference statements)

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“…In order to understand the totipotency/pluripotency of somatic plant cells and to control in vitro regeneration, the genetic factors need to be identified, especially those involved in a specific morphogenic response induced in vitro. The molecular mechanisms induced after the hormone treatment of cultured cells to determine organogenesis or somatic embryogenesis are poorly understood, especially in monocot plants (Kraut et al 2011 ; Su et al 2007 ). A comparison of three transcriptomes in rice cell cultures (somatic embryogenesis, shoot organogenesis, and root organogenesis) pointed to a specific reorganization of the genome-wide transcriptional activities in relation to a particular morphogenic pathway induced in vitro (Su et al 2007 ).…”

Section: Discussionmentioning

confidence: 99%

Morpho-histology and genotype dependence of in vitro morphogenesis in mature embryo cultures of wheat

et al. 2014

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Cellular totipotency is one of the basic principles of plant biotechnology. Currently, the success of the procedure used to produce transgenic plants is directly proportional to the successful insertion of foreign DNA into the genome of suitable target tissue/cells that are able to regenerate plants. The mature embryo (ME) is increasingly recognized as a valuable explant for developing regenerable cell lines in wheat biotechnology. We have previously developed a regeneration procedure based on fragmented ME in vitro culture. Before we can use this regeneration system as a model for molecular studies of the morphogenic pathway induced in vitro and investigate the functional links between regenerative capacity and transformation receptiveness, some questions need to be answered. Plant regeneration from cultured tissues is genetically controlled. Factors such as age/degree of differentiation and physiological conditions affect the response of explants to culture conditions. Plant regeneration in culture can be achieved through embryogenesis or organogenesis. In this paper, the suitability of ME tissues for tissue culture and the chronological series of morphological data observed at the macroscopic level are documented. Genetic variability at each step of the regeneration process was evaluated through a varietal comparison of several elite wheat cultivars. A detailed histological analysis of the chronological sequence of morphological events during ontogeny was conducted. Compared with cultures of immature zygotic embryos, we found that the embryogenic pathway occurs slightly earlier and is of a different origin in our model. Cytological, physiological, and some biochemical aspects of somatic embryo formation in wheat ME culture are discussed.

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Section: Discussionmentioning

confidence: 99%

Morpho-histology and genotype dependence of in vitro morphogenesis in mature embryo cultures of wheat

et al. 2014

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“…The CIM medium contained a basal composition of a B5 medium (Gamborg et al 1968), 0.5 g L − 1 MES, 20 g L − 1 glucose, 2.2 µM of 2,4-D, and 0.2 µM of kinetin (Kraut et al 2011). The SIM-C medium contained micro-elements of MS (Murashige and Skoog 1962), macro-salts, vitamins of a B5 medium (Gamborg et al 1968), and was supplemented with 30 g L − 1 sucrose, 0.5 µM of NAA (1-naphthaleneacetic acid) and 4.4 µM of BAP (6-benzylaminopurine).…”

Section: Methodsmentioning

confidence: 99%

Expression profiling of AUXIN RESPONSE FACTOR genes during somatic embryogenesis induction in Arabidopsis

2017

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Key messageExtensive modulation of numerous ARF transcripts in the embryogenic culture of Arabidopsis indicates a substantial role of auxin signaling in the mechanism of somatic embryogenesis induction.AbstractSomatic embryogenesis (SE) is induced by auxin in plants and auxin signaling is considered to play a key role in the molecular mechanism that controls the embryogenic transition of plant somatic cells. Accordingly, the expression of AUXIN RESPONSE FACTOR (ARF) genes in embryogenic culture of Arabidopsis was analyzed. The study revealed that 14 of the 22 ARFs were transcribed during SE in Arabidopsis. RT-qPCR analysis indicated that the expression of six ARFs (ARF5, ARF6, ARF8, ARF10, ARF16, and ARF17) was significantly up-regulated, whereas five other genes (ARF1, ARF2, ARF3, ARF11, and ARF18) were substantially down-regulated in the SE-induced explants. The activity of ARFs during SE was also monitored with GFP reporter lines and the ARFs that were expressed in areas of the explants engaged in SE induction were detected. A functional test of ARFs transcribed during SE was performed and the embryogenic potential of the arf mutants and overexpressor lines was evaluated. ARFs with a significantly modulated expression during SE coupled with an impaired embryogenic response of the relevant mutant and/or overexpressor line, including ARF1, ARF2, ARF3, ARF5, ARF6, ARF8, and ARF11 were indicated as possibly being involved in SE induction. The study provides evidence that embryogenic induction strongly depends on ARFs, which are key regulators of the auxin signaling. Some clues on the possible functions of the candidate ARFs, especially ARF5, in the mechanism of embryogenic transition are discussed. The results provide guidelines for further research on the auxin-related functional genomics of SE and the developmental plasticity of somatic cells.Electronic supplementary materialThe online version of this article (doi:10.1007/s00299-017-2114-3) contains supplementary material, which is available to authorized users.

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“…The standard CIM medium contained a basal composition of B5 medium (Gamborg et al 1968), 0.5 g L −1 MES, 20 g L −1 glucose, 2.2 μM of 2,4-D and 0.2 μM of kinetin (Feldmann and Marks 1986). Shoot formation in cultures of the CIM-treated seedling roots and IZE-derived cotyledons was induced on SIM-R (Feldmann and Marks 1986) and SIM-C (Kraut et al 2011) regeneration media, respectively. The SIM-R medium was composed of a basal B5 medium (Gamborg et al 1968) and supplemented with 0.5 g L −1 MES, 20 g L −1 glucose, 5 μM of 6-(α,α-dimethylallylamino)-purine (2iP) and 0.8 μM of indole-3-acetic acid (IAA), according to Feldmann and Marks (1986).…”

Section: Methodsmentioning

confidence: 99%

“…The SIM-R medium was composed of a basal B5 medium (Gamborg et al 1968) and supplemented with 0.5 g L −1 MES, 20 g L −1 glucose, 5 μM of 6-(α,α-dimethylallylamino)-purine (2iP) and 0.8 μM of indole-3-acetic acid (IAA), according to Feldmann and Marks (1986). The SIM-C medium contained micro-elements of MS (Murashige and Skoog 1962), macro-salts and vitamins of a B5 medium (Gamborg et al 1968) and was supplemented with 30 g L −1 sucrose, 0.5 μM of NAA and 4.4 μM of BAP (6-benzylaminopurine) (Kraut et al 2011). Modifications of the standard CIM and SIM media included a 10× increase of the auxin concentration to 22 μM 2,4-D in CIM, 8 μM IAA in SIM-R and 5 μM NAA in SIM-C.…”

Section: Methodsmentioning

confidence: 99%

LEAFY COTYLEDON2 (LEC2) promotes embryogenic induction in somatic tissues of Arabidopsis, via YUCCA-mediated auxin biosynthesis

Wójcikowska

Jaskóła

Gąsiorek

et al. 2013

Planta

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138

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The LEAFY COTYLEDON2 (LEC2) transcription factor with a plant-specific B3 domain plays a central role in zygotic and somatic embryogenesis (SE). LEC2 overexpression induced in planta leads to spontaneous somatic embryo formation, but impairs the embryogenic response of explants cultured in vitro under auxin treatment. The auxin-related functions of LEC2 appear during SE induction, and the aim of the present study was to gain further insights into this phenomenon. To this end, the effect of LEC2 overexpression on the morphogenic responses of Arabidopsis explants cultured in vitro under different auxin treatments was evaluated. The expression profiles of the auxin biosynthesis genes were analysed in embryogenic cultures with respect to LEC2 activity. The results showed that LEC2 overexpression severely modifies the requirement of cultured explants for an exogenous auxin concentration at a level that is effective in SE induction and suggested an increase in the auxin content in 35S::LEC2-GR transgenic explants. The assumption of an LEC2 promoted increase in endogenous auxin in cultured explants was further supported by the expression profiling of the genes involved in auxin biosynthesis. The analysis indicated that YUCCAs and TAA1, working in the IPA-YUC auxin biosynthesis pathway, are associated with SE induction, and that the expression of three YUCCA genes (YUC1, YUC4 and YUC10) is associated with LEC2 activity. The results also suggest that the IAOx-mediated auxin biosynthesis pathway involving ATR1/MYB34 and CYP79B2 does not seem to be involved in SE induction. We conclude that de novo auxin production via the tryptophan-dependent IPA-YUC auxin biosynthesis pathway is implicated in SE induction, and that LEC2 plays a key role in this mechanism.Electronic supplementary materialThe online version of this article (doi:10.1007/s00425-013-1892-2) contains supplementary material, which is available to authorized users.

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Immature Zygotic Embryo Cultures of Arabidopsis. A Model System for Molecular Studies on Morphogenic Pathways Induced In Vitro

Cited by 7 publications

References 50 publications

Morpho-histology and genotype dependence of in vitro morphogenesis in mature embryo cultures of wheat

Morpho-histology and genotype dependence of in vitro morphogenesis in mature embryo cultures of wheat

Expression profiling of AUXIN RESPONSE FACTOR genes during somatic embryogenesis induction in Arabidopsis

LEAFY COTYLEDON2 (LEC2) promotes embryogenic induction in somatic tissues of Arabidopsis, via YUCCA-mediated auxin biosynthesis

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