Resistance gene enrichment sequencing (RenSeq) enables reannotation of the NB-LRR gene family from sequenced plant genomes and rapid mapping of resistance loci in segregating populations

RenSeq is a NB-LRR (nucleotide binding-site leucine-rich repeat) gene-targeted, Resistance gene enrichment and sequencing method that enables discovery and annotation of pathogen resistance gene family members in plant genome sequences. We successfully applied RenSeq to the sequenced potato Solanum tuberosum clone DM, and increased the number of identified NB-LRRs from 438 to 755. The majority of these identified R gene loci reside in poorly or previously unannotated regions of the genome. Sequence and positional details on the 12 chromosomes have been established for 704 NB-LRRs and can be accessed through a genome browser that we provide. We compared these NB-LRR genes and the corresponding oligonucleotide baits with the highest sequence similarity and demonstrated that ~80% sequence identity is sufficient for enrichment. Analysis of the sequenced tomato S. lycopersicum ‘Heinz 1706’ extended the NB-LRR complement to 394 loci. We further describe a methodology that applies RenSeq to rapidly identify molecular markers that co-segregate with a pathogen resistance trait of interest. In two independent segregating populations involving the wild Solanum species S. berthaultii (Rpi-ber2) and S. ruiz-ceballosii (Rpi-rzc1), we were able to apply RenSeq successfully to identify markers that co-segregate with resistance towards the late blight pathogen Phytophthora infestans. These SNP identification workflows were designed as easy-to-adapt Galaxy pipelines

A collection of enhancer trap insertional mutants for functional genomics in tomato

[EN] With the completion of genome sequencing projects, the next challenge is to close the gap between gene annotation and gene functional assignment. Genomic tools to identify gene functions are based on the analysis of phenotypic variations between a wild type and its mutant; hence, mutant collections are a valuable resource. In this sense, T-DNA collections allow for an easy and straightforward identification of the tagged gene, serving as the basis of both forward and reverse genetic strategies. This study reports on the phenotypic and molecular characterization of an enhancer trap T-DNA collection in tomato (Solanum lycopersicum L.), which has been produced by Agrobacterium-mediated transformation using a binary vector bearing a minimal promoter fused to the uidA reporter gene. Two genes have been isolated from different T-DNA mutants, one of these genes codes for a UTP-glucose-1-phosphate uridylyltransferase involved in programmed cell death and leaf development, which means a novel gene function reported in tomato. Together, our results support that enhancer trapping is a powerful tool to identify novel genes and regulatory elements in tomato and that this T-DNA mutant collection represents a highly valuable resource for functional analyses in this fleshy-fruited model species.This work was supported by research grants from the Spanish Ministerio de Economia y Competitividad (AGL2012-40150-C02-01, AGL2012-40150-C02-02, AGL2015-64991-C3-1-R and AGL2015-64991-C3-3-R), Junta de Andalucia (P10-AGR-6931) and UE-FEDER. B.P. received a JAE-Doc research contract from the CSIC (Spain). PhD fellowships were funded by the FPU (M.G-A. and R.F.) and the FPI (M.P.A., S.S.A. F-L., A.O-A and L.C.) Programmes of the Ministerio de Ciencia e Innovacion, the JAE predoc Programme of the Spanish CSIC (G.G.), the CONACYT and Universidad de Sinaloa of Mexico (J.S.) and the LASPAU (J.L.Q.). The authors thank research facilities provided by the Campus de Excelencia Internacional Agroalimentario (CeiA3)Pérez-Martín, F.; Yuste-Lisbona, FJ.; Pineda Chaza, BJ.; Angarita-Díaz, MP.; García Sogo, B.; Antón Martínez, MT.; Sanchez Martín-Sauceda, S.... (2017). A collection of enhancer trap insertional mutants for functional genomics in tomato. Plant Biotechnology Journal. 15(11):1439-1452. https://doi.org/10.1111/pbi.12728S14391452151

Mutation at the tomato EXCESSIVE NUMBER OF FLORAL ORGANS (ENO) locus impairs floral meristem development, thus promoting an increased number of floral organs and fruit size

[EN] A novel tomato (Solanum lycopersicum L.) mutant affected in reproductive development, excessive number of floral organs (eno), is described in this study. The eno plants yielded flowers with a higher number of floral organs in the three innermost floral whorls and larger fruits than those found in wild-type plants. Scanning-electron microscopy study indicated that the rise in floral organ number and fruit size correlates with an increased size of floral meristem at early developmental stages. It has been reported that mutation at the FASCIATED (FAS) gene causes the development of flowers with supernumerary organs; however, complementation test and genetic mapping analyses proved that ENO is not an allele of the FAS locus. Furthermore, expression of WUSCHEL (SlWUS) and INHIBITOR OF MERISTEM ACTIVITY (IMA), the two main regulators of floral meristem activity in tomato, is altered in eno but not in fas flowers indicating that ENO could exert its function in the floral meristem independently of FAS. Interestingly, the eno mutation delayed the expression of IMA leading to a prolonged expression of SlWUS, which would explain the greater size of floral meristem. Taken together, results showed that ENO plays a significant role in the genetic pathway regulating tomato floral meristem development. (C) 2014 Elsevier Ireland Ltd. All rights reserved.This work was supported by the research grants AGL2012-40150-C02-01 and AGL2012-40150-C02-02 and a fellowship to AF-L from the Spanish Ministry of Economy and Competitiveness. BP was supported by the European Commission through the JAE-Doc Program of the Spanish National Research Council (CSIC). The authors would like to thank the research facilities provided by the Campus de Excelencia Internacional Agroalimentario (CeiA3).Fernández-Lozano, A.; Yuste-Lisbona, FJ.; Pérez-Martín, F.; Pineda Chaza, BJ.; Moreno Ferrero, V.; Lozano, R.; Angosto, T. (2015). Mutation at the tomato EXCESSIVE NUMBER OF FLORAL ORGANS (ENO) locus impairs floral meristem development, thus promoting an increased number of floral organs and fruit size. Plant Science. 232:41-48. doi:10.1016/j.plantsci.2014.12.007S414823

Досвід застосування лактагелю в практиці акушер-гінеколога

Проведене дослідження показало, що застосування молочної кислоти та глікогену спрямоване на корекцію біохімічних параметрів піхвового середовища (відновлення оптимальних значень pH), сприяє пригніченню росту анаеробних бактерій та створенню оптимальних умов для накопичення власної лактофлори, забезпечуючи колонізаційну резистентність вагінального біотопу та підвищуючи загальну ефективність лікування

The effect of adenosine monophosphate deaminase overexpression on the accumulation of umami-related metabolites in tomatoes

Taste is perceived as one of a combination of five sensations, sweet, sour, bitter, salty, and umami. The umami taste is best known as a savoury sensation and plays a central role in food flavour, palatability, and eating satisfaction. Umami flavour can be imparted by the presence of glutamate and is greatly enhanced by the addition of ribonucleotides, such as inosine monophosphate (IMP) and guanosine monophosphate (GMP). The production of IMP is regulated by the enzyme adenosine monophosphate (AMP) deaminase which functions to convert AMP into IMP. We have generated transgenic tomato (Solanum lycopersicum) lines over expressing AMP deaminase under the control of a fruit-specific promoter. The transgenic lines showed substantially enhanced levels of AMP deaminase expression in comparison to the wild-type control. Elevated AMP deaminase levels resulted in the reduced accumulation of glutamate and increased levels of the umami nucleotide GMP. AMP concentrations were unchanged. The effects on the levels of glutamate and GMP were unexpected and are discussed in relation to the metabolite flux within this pathway

Assessing the genetic variation of Ty-1 and Ty-3 alleles conferring resistance to Tomato Yellow Leaf Curl Virus in a broad tomato germplasm

The online version of this article (doi:10.1007/s11032-015-0329-y) contains supplementary material, which is available to authorized users.[EN] Tomato yellow leaf curl virus (TYLCV) hampers tomato production worldwide. Our previous studies have focussed on mapping and ultimately cloning of the TYLCV resistance genes Ty-1 and Ty-3. Both genes are derived from Solanum chilense and were shown to be allelic. They code for an RNA-dependent RNA polymerase (RDR) belonging to the RDR gamma type defined by a DFDGD catalytic domain. In this study, we first fine-mapped the TYLCV resistance in S. chilense LA1932, LA1960 and LA1971. Results showed that chromosomal intervals of the causal genes in these TYLCV-resistant accessions overlap and cover the region where Ty-1/Ty-3 is located. Further, virus-induced gene silencing was used to silence Ty-1/Ty-3 in tomato lines carrying TYLCV resistance introgressed from S. chilense LA1932, LA1938 and LA1971. Results showed that silencing Ty-1/Ty-3 compromised the resistance in lines derived from S. chilense LA1932 and LA1938. The LA1971-derived material remained resistant upon silencing Ty-1/Ty-3. Further, we studied the allelic variation of the Ty-1/Ty-3 gene by examining cDNA sequences from nine S. chilense-derived lines/accessions and more than 80 tomato cultivars, landraces and accessions of related wild species. The DFDGD catalytic domain of the Ty-1/Ty-3 gene is conserved among all tomato lines and species analysed. In addition, the 12 base pair insertion at the 5-prime part of the Ty-1/Ty-3 gene was found not to be specific for the TYLCV resistance allele. However, compared with the susceptible ty-1 allele, the Ty-1/Ty-3 allele is characterized by three specific amino acids shared by seven TYLCV-resistant S. chilense accessions or derived lines. Thus, Ty-1/Ty-3-specific markers can be developed based on these polymorphisms. Elevated transcript levels were observed for all tested S. chilense RDR alleles (both Ty-1 and ty-1 alleles), demonstrating that elevated expression level is not a good selection criterion for a functional Ty-1/Ty-3 allele.The infectious TYLCV clone was kindly provided by Professor Eduardo Rodriguez Bejarano (Universidad de Malaga). We thank Dick Lohuis for his help with agro-infiltrations, Marc Hendriks and Marjon Arens for RNA isolation and sequencing. This project was financed by the Centre for BioSystems Genomics (CBSG), which is part of the Netherlands Genomics Initiative/Netherlands Organization for Scientific Research (http://www.cbsg.nl). Olga Julian was granted a scholarship by Generalitat Valenciana. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.Caro, M.; Verlaan, MG.; Julián Rodríguez, O.; Finkers, R.; Wolters, AA.; Hutton, S.; Scott, JW.... (2015). Assessing the genetic variation of Ty-1 and Ty-3 alleles conferring resistance to Tomato Yellow Leaf Curl Virus in a broad tomato germplasm. Molecular Breeding. 35. doi:10.1007/s11032-015-0329-yS13235Agrama H, Scott J (2006) Quantitative trait loci for tomato yellow leaf curl virus and tomato mottle virus resistance in tomato. 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The tomato res mutant which accumulates JA in roots in non-stressed conditions restores cell structure alterations under salinity

[EN] Jasmonic acid (JA) regulates a wide spectrum of plant biological processes, from plant development to stress defense responses. The role of JA in plant response to salt stress is scarcely known, and even less known is the specific response in root, the main plant organ responsible for ionic uptake and transport to the shoot. Here we report the characterization of the first tomato (Solanum lycopersicum)mutant, named res (restored cell structure by salinity), that accumulates JA in roots prior to exposure to stress. The res tomato mutant presented remarkable growth inhibition and displayed important morphological alterations and cellular disorganization in roots and leaves under control conditions, while these alterations disappeared when the res mutant plants were grown under salt stress. Reciprocal grafting between res and wild type (WT) (tomato cv. Moneymaker) indicated that the main organ responsible for the development of alterations was the root. The JA-signaling pathway is activated in res roots prior to stress, with transcripts levels being even higher in control condition than in salinity. Future studies on this mutant will provide significant advances in the knowledge of JA role in root in salt-stress tolerance response, as well as in the energy trade-off between plant growth and response to stress. Guardar / Salir Siguiente >This work was supported by grant AGL2012-40150-C03 from the Spanish 'Ministerio de Economia'; The Spanish Council of Scientific Research (CSIC) [postdoctoral position from the JAE-Doc programme grant E-30-2011-0443170 conceded to IE and BP]; Spanish 'Ministerio de Economia' [postdoctoral position from the RyC programme grant RYC2010-06369 conceded to NFG].García-Abellán, JO.; Fernández-García, N.; López-Berenguer, C.; Egea, I.; Flores, FB.; Angosto, T.; Capel, J.... (2015). The tomato res mutant which accumulates JA in roots in non-stressed conditions restores cell structure alterations under salinity. Physiologia Plantarum. 155(3):296-314. https://doi.org/10.1111/ppl.12320S296314155