Transposon Tagging of the Defective embryo and meristems Gene of Tomato

Keddie, James S.; Carroll, Bernard J.; Reyes, M.; Klimyuk, Victor; Holtan, Hans; Gruissem, Wilhelm; Jones, Jonathan D. G.

doi:10.2307/3870675

Cited by 10 publications

(12 citation statements)

References 22 publications

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“…Finally, the development of an effective progeny-screening protocol makes this population and its derivatives capable of functioning in a high-throughput system. These factors make the transgenic population developed during the course of this project a unique, new resource for functional genomics of tomato with a new array of phenotypic mutants available to augment those that have already been characterized from earlier transposon-tagging schemes (Bishop et al, 1996;van der Biezen et al, 1996;Keddie et al, 1998).…”

Section: Discussionmentioning

confidence: 99%

An ActiveAc/DsTransposon System for Activation Tagging in Tomato Cultivar M82 Using Clonal Propagation

et al. 2013

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Tomato (Solanum lycopersicum) is a model organism for Solanaceae in both molecular and agronomic research. This project utilized Agrobacterium tumefaciens transformation and the transposon-tagging construct Activator (Ac)/Dissociator (Ds)-ATag-Bar_gosGFP to produce activation-tagged and knockout mutants in the processing tomato cultivar M82. The construct carried hygromycin resistance (hyg), green fluorescent protein (GFP), and the transposase (TPase) of maize (Zea mays) Activator major transcript X054214.1 on the stable Ac element, along with a 35S enhancer tetramer and glufosinate herbicide resistance (BAR) on the mobile Ds-ATag element. An in vitro propagation strategy was used to produce a population of 25 T0 plants from a single transformed plant regenerated in tissue culture. A T1 population of 11,000 selfed and cv M82 backcrossed progeny was produced from the functional T0 line. This population was screened using glufosinate herbicide, hygromycin leaf painting, and multiplex polymerase chain reaction (PCR). Insertion sites of transposed Ds-ATag elements were identified through thermal asymmetric interlaced PCR, and resulting product sequences were aligned to the recently published tomato genome. A population of 509 independent, Ds-only transposant lines spanning all 12 tomato chromosomes has been developed. Insertion site analysis demonstrated that more than 80% of these lines harbored Ds insertions conducive to activation tagging. The capacity of the Ds-ATag element to alter transcription was verified by quantitative real-time reverse transcription-PCR in two mutant lines. The transposon-tagged lines have been immortalized in seed stocks and can be accessed through an online database, providing a unique resource for tomato breeding and analysis of gene function in the background of a commercial tomato cultivar.

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Section: Discussionmentioning

confidence: 99%

An ActiveAc/DsTransposon System for Activation Tagging in Tomato Cultivar M82 Using Clonal Propagation

et al. 2013

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“…These include the Pto kinase (12) and the closely related Fen kinase from tomato (13), the ATN1 (14) and APK1 (15) kinases from Arabidopsis, several calcium-dependent kinases (CDPKs) from maize (16), rice (17), tobacco (18), and Arabidopsis (19), calcineurin ␤-like proteins from Arabidopsis (20), and G␣ subunits of the Gproteins from several plant species (21). A myristoylation motif is also found in the predicted amino acid sequences of fatty acid acyl-CoA synthetase gene from Brassica napus (22), a nodulespecific gene family from Alnus glutinosa (23), and the Dem (defective embryo and meristems) gene from tomato (24). These observations imply that protein N-myristoylation may be involved in various plant processes.…”

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confidence: 93%

“…In fact, a number of plant proteins contain putative consensus myristoylation motifs (MGXXX(S/T)-, Refs. [12][13][14][15][16][17][18][19][20][21][22][23][24]. These include the Pto kinase (12) and the closely related Fen kinase from tomato (13), the ATN1 (14) and APK1 (15) kinases from Arabidopsis, several calcium-dependent kinases (CDPKs) from maize (16), rice (17), tobacco (18), and Arabidopsis (19), calcineurin ␤-like proteins from Arabidopsis (20), and G␣ subunits of the Gproteins from several plant species (21).…”

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confidence: 99%

Molecular Cloning, Genomic Organization, and Biochemical Characterization of Myristoyl-CoA:ProteinN-Myristoyltransferase from Arabidopsis thaliana

Qi¹,

Rajala²,

Anderson³

et al. 2000

Journal of Biological Chemistry

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Myristoyl-CoA:protein N-myristoyltransferase (NMT, EC 2.3.1.97) catalyzes the co-translational addition of myristic acid to the amino-terminal glycine residue of a number of important proteins of diverse functions. We have isolated a full-length Arabidopsis thaliana cDNA encoding NMT (AtNMT1), the first described from a higher plant. This AtNMT1 cDNA clone has an open reading frame of 434 amino acids and a predicted molecular mass of 48,706 Da. The primary structure is 50% identical to the mammalian NMTs. Analyses of Southern blots, genomic clones, and database sequences suggested that the A. thaliana genome contains two copies of NMT gene, which are present on different chromosomes and have distinct genomic organizations. The recombinant AtNMT1 expressed in Escherichia coli exhibited a high catalytic efficiency for the peptides derived from putative plant myristoylated proteins AtCDPK6 and Fen kinase. The AtNMT was similar to the mammalian NMTs with respect to a relative specificity for myristoyl CoA amoung the acyl CoA donors and also inhibition by the bovine brain NMT inhibitor NIP 71 . The AtNMT1 expression profile indicated ubiquity in roots, stem, leaves, flowers, and siliques (Ϸ1.7 kb transcript and Ϸ50 kDa immunoreactive polypeptide) but a greater level in the younger tissue, which are developmentally very active. NMT activity was also evident in all these tissues. Subcellular distribution studies indicated that, in leaf extracts, ϳ60% of AtNMT activity was associated with the ribosomal fractions, whereas ϳ30% of the activity was observed in the cytosolic fractions. The NMT is biologically important to plants, as noted from the stunted development when the AtNMT1 was down-regulated in transgenic Arabidopsis under the control of an enhanced CaMV 35S promoter. The results presented in this study provide the first direct molecular evidence for plant protein N-myristoylation and a mechanistic basis for understanding the role of this protein modification in plants.

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“…One line was named poc1-1 and backcrossed twice for further characterization. The crossing of poc mutants with the Ds transposontagged dem mutant (Keddie et al, 1998), which also has a variable number of cotyledons, revealed lack of allellism between these two mutants (M. Kavitha, M.S. Sharada, P. Janila, S. Negi, R. Sharma, unpublished data).…”

Section: Genetic Characterization Of the Polycot Mutantmentioning

confidence: 99%

The polycotyledon Mutant of Tomato Shows Enhanced Polar Auxin Transport

et al. 2003

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The polycotyledon mutant of tomato (Lycopersicon esculentum L. cv Ailsa Craig) showed altered development during embryogenesis and during vegetative and reproductive phases. The phenotype was pleiotropic and included the formation of extra cotyledons, changes in leaf shape, increased number of flowers (indeterminacy) with abnormal floral organs, the formation of epiphyllous structures, and altered gravitropism. The earliest defects were observed at the transition from the globular to the heart stage of embryogenesis with the formation of multiple cotyledons. Epidermal cells in the mutant embryo were smaller and less expanded compared with wild type. Examination of polar auxin transport (PAT) showed a striking enhancement in the case of the mutant. Increase in PAT did not appear to be caused by a decrease in flavonoids because the mutant had normal flavonoid levels. Application of 2,3,5-triiodobenzoic acid, an inhibitor of polar transport of auxin, rescued postgermination phenotypes of young seedlings. Our analysis reveals a level of control that negatively regulates PAT in tomato and its contribution to plant development and organogenesis.

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Transposon Tagging of the Defective embryo and meristems Gene of Tomato

Cited by 10 publications

References 22 publications

An ActiveAc/DsTransposon System for Activation Tagging in Tomato Cultivar M82 Using Clonal Propagation

An ActiveAc/DsTransposon System for Activation Tagging in Tomato Cultivar M82 Using Clonal Propagation

Molecular Cloning, Genomic Organization, and Biochemical Characterization of Myristoyl-CoA:ProteinN-Myristoyltransferase from Arabidopsis thaliana

The polycotyledon Mutant of Tomato Shows Enhanced Polar Auxin Transport

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