Optimizing TILLING populations for reverse genetics in <i>Medicago truncatula</i>

Signor, Christine Le; Savois, Vincent; Aubert, Grégoire; Verdier, Jérôme; Nicolas, Marie; Pagny, Gaëlle; Moussy, F.; Sánchez, Myriam; Baker, D. M.; Clarke, Jonathan H.; Thompson, Richard

doi:10.1111/j.1467-7652.2009.00410.x

Cited by 96 publications

(66 citation statements)

References 26 publications

(39 reference statements)

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“…Firstly, a TILLING approach was employed to identify mutations within a 1 kb region of the ERN2 coding sequence by screening an ethyl methanesulfonate (EMS)-mutagenized collection (Le Signor et al, 2009). A single-nucleotide substitution mutant, designated ern2-1, was shown to carry a threonine (Thr, T) to isoleucine (Ile, I) transition at position 61 within the ERN2 DNA binding domain (Fig.…”

Section: Identification Of Two Ern2 Mutant Allelesmentioning

confidence: 99%

“…Despite the fact that the two ern2 lines were isolated from A17 and R108 mutant collections (Le Signor et al, 2009;Pislariu et al, 2012), they display similar phenotypes. Although forming functional nitrogen-fixing nodules, ern2 mutant lines nevertheless exhibit some limited nodulation defects since we observe an overall 2-fold decrease in the number of infected nodules and nodule primordia as well as certain nodules containing a limited number of abnormal ITs (see Fig.…”

Section: Mild Infection Defects May Contribute To the Accelerated Senmentioning

confidence: 99%

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The Symbiosis-Related ERN Transcription Factors Act in Concert to Coordinate Rhizobial Host Root Infection

et al. 2016

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Legumes improve their mineral nutrition through nitrogen-fixing root nodule symbioses with soil rhizobia. Rhizobial infection of legumes is regulated by a number of transcription factors, including ERF Required for Nodulation1 (ERN1). Medicago truncatula plants defective in ERN1 are unable to nodulate, but still exhibit early symbiotic responses including rhizobial infection. ERN1 has a close homolog, ERN2, which shows partially overlapping expression patterns. Here we show that ern2 mutants exhibit a later nodulation phenotype than ern1, being able to form nodules but with signs of premature senescence. Molecular characterization of the ern2-1 mutation reveals a key role for a conserved threonine for both DNA binding and transcriptional activity. In contrast to either single mutant, the double ern1-1 ern2-1 line is completely unable to initiate infection or nodule development. The strong ern1-1 ern2-1 phenotype demonstrates functional redundancy between these two transcriptional regulators and reveals the essential role of ERN1/ERN2 to coordinately induce rhizobial infection and nodule organogenesis. While ERN1/ERN2 act in concert in the root epidermis, only ERN1 can efficiently allow the development of mature nodules in the cortex, probably through an independent pathway. Together, these findings reveal the key roles that ERN1/ERN2 play at the very earliest stages of root nodule development.

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Section: Identification Of Two Ern2 Mutant Allelesmentioning

confidence: 99%

Section: Mild Infection Defects May Contribute To the Accelerated Senmentioning

confidence: 99%

The Symbiosis-Related ERN Transcription Factors Act in Concert to Coordinate Rhizobial Host Root Infection

et al. 2016

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“…TILLING on the genes LjPGM1, LjAPS1, LjAPL2, and LjGWD1 was performed as described previously (Horst et al, 2007) using gene-specific primers. TILLING for LjGWD3 was carried out by RevGenUK (http://revgenuk.jic.ac.uk/) on an ABI3730 capillary sequencer following the method of Le Signor et al (2009). Gene-specific primers used for TILLING are shown in Supplemental Table S4.…”

Section: Tillingmentioning

confidence: 99%

A Suite of Lotus japonicus Starch Mutants Reveals Both Conserved and Novel Features of Starch Metabolism

et al. 2010

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The metabolism of starch is of central importance for many aspects of plant growth and development. Information on leaf starch metabolism other than in Arabidopsis (Arabidopsis thaliana) is scarce. Furthermore, its importance in several agronomically important traits exemplified by legumes remains to be investigated. To address this issue, we have provided detailed information on the genes involved in starch metabolism in Lotus japonicus and have characterized a comprehensive collection of forward and TILLING (for Targeting Induced Local Lesions IN Genomes) reverse genetics mutants affecting five enzymes of starch synthesis and two enzymes of starch degradation. The mutants provide new insights into the structure-function relationships of ADP-glucose pyrophosphorylase and glucan, water dikinase1 in particular. Analyses of the mutant phenotypes indicate that the pathways of leaf starch metabolism in L. japonicus and Arabidopsis are largely conserved. However, the importance of these pathways for plant growth and development differs substantially between the two species. Whereas essentially starchless Arabidopsis plants lacking plastidial phosphoglucomutase grow slowly relative to wild-type plants, the equivalent mutant of L. japonicus grows normally even in a 12-h photoperiod. In contrast, the loss of GLUCAN, WATER DIKINASE1, required for starch degradation, has a far greater effect on plant growth and fertility in L. japonicus than in Arabidopsis. Moreover, we have also identified several mutants likely to be affected in new components or regulators of the pathways of starch metabolism. This suite of mutants provides a substantial new resource for further investigations of the partitioning of carbon and its importance for symbiotic nitrogen fixation, legume seed development, and perenniality and vegetative regrowth.Recent studies in Arabidopsis (Arabidopsis thaliana) have greatly enhanced our knowledge about pathways of transitory starch metabolism (Zeeman et al., 2007;Keeling and Myers, 2010;Kö tting et al., 2010;Zeeman et al., 2010). The pathway of synthesis is well established for several species, but the degradative pathway is understood only in Arabidopsis. During synthesis, the plastidial isoforms of phosphoglucoisomerase (PGI1) and phosphoglucomutase (PGM1), together with ADP-Glc pyrophosphorylase (AGPase), catalyze the conversion of the Calvin cycle intermediate Fru 6-P to ADPGlc, the substrate for starch synthases (Supplemental Fig. S1). Leaves of mutants lacking any of these three enzymes either have strongly reduced starch contents or lack starch almost completely (Caspar et al., 1985;Hanson and McHale, 1988;Lin et al., 1988aLin et al., , 1988bKruckeberg et al., 1989;Harrison et al., 1998;Yu et al., 2000;Streb et al., 2009). In contrast, the phenotypes of mutants lacking individual enzymes that convert ADPGlc into starch vary between species and are often much less pronounced (starch synthases [Delvallé et al., 2005;Zhang et al., 2005] and starch-branching enzymes [Tomlinson et al., 1997;Blauth et al....

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“…However, systematic mutant populations have been developed in several species in order to address this issue. TILLING populations are available for L. japonicus, pea, M. truncatula and chickpea (Dalmais et al, 2008;Le-Signor et al, 2009;Perry et al, 2003;Varshney et al, unpublished), Fast Neutron generated deletion mutant populations for M. truncatula, pea and soybean (Bolon et al, 2011;Hofer et al, 2009;Rogers et al, 2009;Smykal et al, 2012) and insertion mutant populations generated using exogenous (Medicago, d 'Erfurth et al, 2003) and endogeneous retrotransposons (Lotus, Urbański et al, 2012) have been generated and have contributed to gene discovery in these species (see Domoney et al, 2013;Domonkos et al, 2013;Urbański et al, 2013).…”

Section: Glyma07g05410/ Glyma16g01980mentioning

confidence: 99%