| 1. |
- Pacurar, Daniel I, et al.
(författare)
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Novel viable allele of Arabidopsis CULLIN1 identified in a screen for superroot2 suppressors by next generation sequencing-assisted mapping
- 2014
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Ingår i: PLOS ONE. - : Public library science. - 1932-6203. ; 9:6, s. e100846-
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Tidskriftsartikel (refereegranskat)abstract
- Map-based cloning (MBC) is the conventional approach for linking phenotypes to genotypes, and has been successfully used to identify causal mutations in diverse organisms. Next-generation sequencing (NGS) technologies offer unprecedented possibilities to sequence the entire genomes of organisms, thereby in principle enabling direct identification of causal mutations without mapping. However, although mapping-by-sequencing has proven to be a cost effective alternative to classical MBC in particular situations, methods based solely on NGS still have limitations and need to be refined. Aiming to identify the causal mutations in suppressors of Arabidopsis thaliana superroot2 phenotype, generated by ethyl methane sulfonate (EMS) treatment, we combined NGS and classical mapping, to rapidly identify the point mutations and restrict the number of testable candidates by defining the chromosomal intervals containing the causal mutations, respectively. The NGS-assisted mapping approach we describe here facilitates unbiased identification of virtually any causal EMS-generated mutation by overlapping the identification (deep sequencing) and validation (mapping) steps. To exemplify the useful marriage of the two approaches we discuss the strategy used to identify a new viable recessive allele of the Arabidopsis CULLIN1 gene in the non-reference Wassilewskija (Ws-4) accession.
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| 2. |
- Pacurar, Daniel Ioan, et al.
(författare)
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The Arabidopsis Cop9 signalosome subunit 4 (CNS4) is involved in adventitious root formation
- 2017
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Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 7
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Tidskriftsartikel (refereegranskat)abstract
- The COP9 signalosome (CSN) is an evolutionary conserved multiprotein complex that regulates many aspects of plant development by controlling the activity of CULLIN-RING E3 ubiquitin ligases (CRLs). CRLs ubiquitinate and target for proteasomal degradation a vast number of specific substrate proteins involved in many developmental and physiological processes, including light and hormone signaling and cell division. As a consequence of CSN pleiotropic function, complete loss of CSN activity results in seedling lethality. Therefore, a detailed analysis of CSN physiological functions in adult Arabidopsis plants has been hampered by the early seedling lethality of csn null mutants. Here we report the identification and characterization of a viable allele of the Arabidopsis COP9 signalosome subunit 4 (CSN4). The allele, designated csn4-2035, suppresses the adventitious root (AR) phenotype of the Arabidopsis superroot2-1 mutant, potentially by altering its auxin signaling. Furthermore, we show that although the csn4-2035 mutation affects primary and lateral root (LR) formation in the 2035 suppressor mutant, CSN4 and other subunits of the COP9 complex seem to differentially control AR and LR development.
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| 3. |
- Gutierrez, Laurent, et al.
(författare)
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Auxin Controls Arabidopsis Adventitious Root Initiation by Regulating Jasmonic Acid Homeostasis
- 2012
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Ingår i: The Plant Cell. - : Oxford University Press (OUP). - 1040-4651 .- 1532-298X. ; 24:6, s. 2515-2527
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Tidskriftsartikel (refereegranskat)abstract
- Vegetative shoot-based propagation of plants, including mass propagation of elite genotypes, is dependent on the development of shoot-borne roots, which are also called adventitious roots. Multiple endogenous and environmental factors control the complex process of adventitious rooting. In the past few years, we have shown that the auxin response factors ARF6 and ARF8, targets of the microRNA miR167, are positive regulators of adventitious rooting, whereas ARF17, a target of miR160, is a negative regulator. We showed that these genes have overlapping expression profiles during adventitious rooting and that they regulate each other's expression at the transcriptional and posttranscriptional levels by modulating the homeostasis of miR160 and miR167. We demonstrate here that this complex network of transcription factors regulates the expression of three auxin-inducible Gretchen Hagen3 (GH3) genes, GH3.3, GH3.5, and GH3.6, encoding acyl-acid-amido synthetases. We show that these three GH3 genes are required for fine-tuning adventitious root initiation in the Arabidopsis thaliana hypocotyl, and we demonstrate that they act by modulating jasmonic acid homeostasis. We propose a model in which adventitious rooting is an adaptive developmental response involving crosstalk between the auxin and jasmonate regulatory pathways.
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| 4. |
- Gutierrez, Laurent, et al.
(författare)
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Phenotypic plasticity of adventitious rooting in Arabidopsis is controlled by complex regulation of Auxin response factor transcripts and microRNA abundance
- 2009
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Ingår i: The Plant Cell. - : Oxford University Press (OUP). - 1040-4651 .- 1532-298X. ; 21:10, s. 3119-3132
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Tidskriftsartikel (refereegranskat)abstract
- The development of shoot-borne roots, or adventitious roots, is indispensable for mass propagation of elite genotypes. It is a complex genetic trait with a high phenotypic plasticity due to multiple endogenous and environmental regulatory factors. We demonstrate here that a subtle balance of activator and repressor AUXIN RESPONSE FACTOR (ARF) transcripts controls adventitious root initiation. Moreover, microRNA activity appears to be required for fine-tuning of this process. Thus, ARF17, a target of miR160, is a negative regulator, and ARF6 and ARF8, targets of miR167, are positive regulators of adventitious rooting. The three ARFs display overlapping expression domains, interact genetically, and regulate each other's expression at both transcriptional and posttranscriptional levels by modulating miR160 and miR167 availability. This complex regulatory network includes an unexpected feedback regulation of microRNA homeostasis by direct and nondirect target transcription factors. These results provide evidence of microRNA control of phenotypic variability and are a significant step forward in understanding the molecular mechanisms regulating adventitious rooting.
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| 5. |
- Pacurar, Daniel Ioan, et al.
(författare)
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A collection of INDEL markers for map-based cloning in seven Arabidopsis accessions
- 2012
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Ingår i: Journal of Experimental Botany. - : Oxford University Press (OUP). - 0022-0957 .- 1460-2431. ; 63:7, s. 2491-2501
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Tidskriftsartikel (refereegranskat)abstract
- The availability of a comprehensive set of resources including an entire annotated reference genome, sequenced alternative accessions, and a multitude of marker systems makes Arabidopsis thaliana an ideal platform for genetic mapping. PCR markers based on INsertions/DELetions (INDELs) are currently the most frequently used polymorphisms. For the most commonly used mapping combination, ColumbiaxLandsberg erecta (Col-0xLer-0), the Cereon polymorphism database is a valuable resource for the generation of polymorphic markers. However, because the number of markers available in public databases for accessions other than Col-0 and Ler-0 is extremely low, mapping using other accessions is far from straightforward. This issue arose while cloning mutations in the Wassilewskija (Ws-4) background. In this work, approaches are described for marker generation in Ws-4 x Col-0. Complementary strategies were employed to generate 229 INDEL markers. Firstly, existing Col-0/Ler-0 Cereon predicted polymorphisms were mined for transferability to Ws-4. Secondly, Ws-0 ecotype Illumina sequence data were analyzed to identify INDELs that could be used for the development of PCR-based markers for Col-0 and Ws-4. Finally, shotgun sequencing allowed the identification of INDELs directly between Col-0 and Ws-4. The polymorphism of the 229 markers was assessed in seven widely used Arabidopsis accessions, and PCR markers that allow a clear distinction between the diverged Ws-0 and Ws-4 accessions are detailed. The utility of the markers was demonstrated by mapping more than 35 mutations in a Col-0xWs-4 combination, an example of which is presented here. The potential contribution of next generation sequencing technologies to more traditional map-based cloning is discussed.
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| 6. |
- Pacurar, Daniel I., et al.
(författare)
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Agrobacterium tumefaciens : From crown gall tumors to genetic transformation
- 2011
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Ingår i: Physiological and molecular plant pathology. - London : Academic Press. - 0885-5765 .- 1096-1178. ; 76:2, s. 76-81
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Forskningsöversikt (refereegranskat)abstract
- The phytopathogenic bacterium Agrobacterium tumefaciens is the causative agent of crown gall disease on a broad range of plant species. For more than a century this pathogen has fascinated biologists for various reasons. It has been believed that disclosing the mystery of the crown gall tumour disease in plants might help to understand the mechanisms of oncogenesis in general, and eventually use this knowledge to cure cancer in animals and humans. However, as the performer of the only known case of trans-kingdom DNA transfer, this natural genetic engineer is recognised by nowadays biologists particularly for this feature, and Agrobacterium-mediated genetic transformation has become the preferred method to generate transgenic plants. This review briefly highlights the key steps in DNA transfer during natural A. tumefaciens infection and the subsequent development of crown gall disease. It will focus on the adaptation and use of this proof of principle for the purpose of genetic transformation. (C) 2011 Elsevier Ltd. All rights reserved.
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| 7. |
- Pacurar, Daniel Ioan, et al.
(författare)
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Identification of new adventitious rooting mutants amongst suppressors of the Arabidopsis thaliana superroot2 mutation
- 2014
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Ingår i: Journal of Experimental Botany. - : Oxford University Press. - 0022-0957 .- 1460-2431. ; 65:6, s. 1605-1618
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Tidskriftsartikel (refereegranskat)abstract
- The plant hormone auxin plays a central role in adventitious rooting and is routinely used with many economically important, vegetatively propagated plant species to promote adventitious root initiation and development on cuttings. Nevertheless the molecular mechanisms through which it acts are only starting to emerge. The Arabidopsis superroot2-1 (sur2-1) mutant overproduces auxin and, as a consequence, develops excessive adventitious roots in the hypocotyl. In order to increase the knowledge of adventitious rooting and of auxin signalling pathways and crosstalk, this study performed a screen for suppressors of superroot2-1 phenotype. These suppressors provide a new resource for discovery of genetic players involved in auxin signalling pathways or at the crosstalk of auxin and other hormones or environmental signals. This study reports the identification and characterization of 26 sur2-1 suppressor mutants, several of which were identified as mutations in candidate genes involved in either auxin biosynthesis or signalling. In addition to confirming the role of auxin as a central regulator of adventitious rooting, superroot2 suppressors indicated possible crosstalk with ethylene signalling in this process.
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