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

Identifying Molecular Markers Suitable For Frl Selection in Tomato Breeding

Modern plant breeding heavily relies on the use of molecular markers. In recent years, next generation sequencing (NGS) emerged as a powerful technology to discover DNA sequence polymorphisms and generate molecular markers very rapidly and cost effectively, accelerating the plant breeding programmes. A single dominant locus, Frl, in tomato provides resistance to the fungal pathogen Fusarium oxysporum f. sp. radicis-lycopersici (FORL), causative agent of Fusarium crown and root rot. In this study, we describe the generation of molecular markers associated with the Frl locus. An F2 mapping population between an FORL resistant and a susceptible cultivar was generated. NGS technology was then used to sequence the genomes of a susceptible and a resistant parent as well the genomes of bulked resistant and susceptible F2 lines. We zoomed into the Frl locus and mapped the locus to a 900 kb interval on chromosome 9. Polymorphic single-nucleotide polymorphisms (SNPs) within the interval were identified and markers co-segregating with the resistant phenotype were generated. Some of these markers were tested successfully with commercial tomato varieties indicating that they can be used for marker-assisted selection in large-scale breeding programmes

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