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Mariner transposon

Yongming Wang, Diana Pryputniewicz-Dobrinska, Enikö Éva Nagy, Christopher D Kaufman, Manvendra Singh, Steve Yant, Jichang Wang, Anna Dalda, Mark A Kay, Zoltán Ivics, Zsuzsanna Izsvák
The functional relevance of the inverted repeat structure (IR/DR) in a subgroup of the Tc1/mariner superfamily of transposons has been enigmatic. In contrast to mariner transposition, where a topological filter suppresses single-ended reactions, the IR/DR orchestrates a regulatory mechanism to enforce synapsis of the transposon ends before cleavage by the transposase occurs. This ordered assembly process shepherds primary transposase binding to the inner 12DRs (where cleavage does not occur), followed by capture of the 12DR of the other transposon end...
December 1, 2016: Nucleic Acids Research
Natalia Guschinskaya, Romain Brunel, Maxime Tourte, Gina L Lipscomb, Michael W W Adams, Philippe Oger, Xavier Charpentier
Transposition mutagenesis is a powerful tool to identify the function of genes, reveal essential genes and generally to unravel the genetic basis of living organisms. However, transposon-mediated mutagenesis has only been successfully applied to a limited number of archaeal species and has never been reported in Thermococcales. Here, we report random insertion mutagenesis in the hyperthermophilic archaeon Pyrococcus furiosus. The strategy takes advantage of the natural transformability of derivatives of the P...
November 8, 2016: Scientific Reports
Maelin da Silva, Patricia Barbosa, Roberto F Artoni, Eliana Feldberg
Gymnotidae is a family of electric fish endemic to the Neotropics consisting of 2 genera: Electrophorus and Gymnotus. The genus Gymnotus is widely distributed and is found in all of the major Brazilian river systems. Physical and molecular mapping data for the ribosomal DNA (rDNA) in this genus are still scarce, with its chromosomal location known in only 11 species. As other species of Gymnotus with 2n = 54 chromosomes from the Paraná-Paraguay basin, G. mamiraua was found to have a large number of 5S rDNA sites...
October 18, 2016: Cytogenetic and Genome Research
Mai Yamamoto, Takashige Kashimoto, Yukihiro Yoshimura, Nao Tachibana, Shiho Kuroda, Yoshiko Miki, Sou Kitabayashi, Ping Tong, Jianbo Xiao, Koichi Tanaka, Hiroshi Hamamoto, Kazuhisa Sekimizu, Koichiro Yamamoto
The halophilic marine bacterium, Vibrio vulnificus, occasionally causes fatal septicemia in immunocompromised patients. Mice are commonly used as experimental animals to investigate the virulence of V. vulnificus, however, a large number of mice are generally required for bioassays. The present study examined whether the invertebrate species, silkworms, can be used instead of mice to investigate V. vulnificus virulence. When the silkworms were inoculated with 1.2x107 colony forming units of V. vulnificus OPU1‑Rf, a virulent strain of V...
September 26, 2016: Molecular Medicine Reports
Brian J Akerley
The property of transposons to randomly insert into target DNA has long been exploited for generalized mutagenesis and forward genetic screens. Newer applications that monitor the relative abundance of each transposon insertion in large libraries of mutants can be used to evaluate the roles in cellular fitness of all genes of an organism, provided that transposition is in fact random across all genes. In a recent article, Kimura and colleagues identified an important exception to the latter assumption [S. Kimura, T...
October 11, 2016: MBio
Nengding Wang, Egon A Ozer
Transposon insertion sequencing is a process whereby microbial fitness determinants can be identified on a genome-wide scale. This process uses high-throughput next generation sequencing to screen for changes in the composition of a pool of transposon mutants after exposure to selective conditions. One commonly used process for generating transposon insertion sequencing libraries is called INSeq that works with mutant pools produced using a modified Mariner transposon. Libraries produced using the INSeq process are sequenced on the Illumina platform...
2017: Methods in Molecular Biology
Mariane S Thøgersen, Marina W Delpin, Jette Melchiorsen, Mogens Kilstrup, Maria Månsson, Boyke Bunk, Cathrin Spröer, Jörg Overmann, Kristian F Nielsen, Lone Gram
It has previously been reported that some strains of the marine bacterium Pseudoalteromonas luteoviolacea produce the purple bioactive pigment violacein as well as the antibiotic compound indolmycin, hitherto only found in Streptomyces. The purpose of the present study was to determine the relative role of each of these two compounds as antibacterial compounds in P. luteoviolacea S4054. Using Tn10 transposon mutagenesis, a mutant strain that was significantly reduced in violacein production in mannose-containing substrates was created...
2016: Frontiers in Microbiology
Federico Rosconi, Stefan P W de Vries, Abiyad Baig, Elena Fabiano, Andrew J Grant
: The interior of plants contains microorganisms (referred to as endophytes) distinct to those present in the root surface or in the surrounding soil. Herbaspirillum seropedicae strain SmR1, belonging to the β-Proteobacterium, is an endophyte that colonizes crops, including rice, maize, sugarcane and sorghum. Different approaches have revealed genes and pathways regulated during the interaction of H. seropedicae with its plant hosts. However, functional genomic analysis of transposon (Tn) mutants has been hampered by the lack of genetic tools...
September 2, 2016: Applied and Environmental Microbiology
Satoshi Kimura, Troy P Hubbard, Brigid M Davis, Matthew K Waldor
UNLABELLED: Transposon insertion sequencing (TIS; also known as TnSeq) is a potent approach commonly used to comprehensively define the genetic loci that contribute to bacterial fitness in diverse environments. A key presumption underlying analyses of TIS datasets is that loci with a low frequency of transposon insertions contribute to fitness. However, it is not known whether factors such as nucleoid binding proteins can alter the frequency of transposon insertion and thus whether TIS output may systematically reflect factors that are independent of the role of the loci in fitness...
2016: MBio
Michelle Orane Schemberger, Viviane Nogaroto, Mara Cristina Almeida, Roberto Ferreira Artoni, Guilherme Targino Valente, Cesar Martins, Orlando Moreira-Filho, Marta Margarete Cestari, Marcelo Ricardo Vicari
Transposable elements are able to move along eukaryotic genomes. They are divided into two classes according to their transposition intermediate: RNA (class I or retrotransposons) or DNA (class II or DNA transposons). Most of these sequences are inactive or non-autonomous in eukaryotic genomes. Inactivate transposons can accumulate mutations at neutral rates until losing their molecular identity. They may either be eliminated from the genome or take on different molecular functions. Transposable elements may also participate in the differentiation of sex chromosomes...
November 30, 2016: Gene
Partow Kebriaei, Harjeet Singh, M Helen Huls, Matthew J Figliola, Roland Bassett, Simon Olivares, Bipulendu Jena, Margaret J Dawson, Pappanaicken R Kumaresan, Shihuang Su, Sourindra Maiti, Jianliang Dai, Branden Moriarity, Marie-Andrée Forget, Vladimir Senyukov, Aaron Orozco, Tingting Liu, Jessica McCarty, Rineka N Jackson, Judy S Moyes, Gabriela Rondon, Muzaffar Qazilbash, Stefan Ciurea, Amin Alousi, Yago Nieto, Katy Rezvani, David Marin, Uday Popat, Chitra Hosing, Elizabeth J Shpall, Hagop Kantarjian, Michael Keating, William Wierda, Kim Anh Do, David A Largaespada, Dean A Lee, Perry B Hackett, Richard E Champlin, Laurence J N Cooper
BACKGROUND: T cells expressing antigen-specific chimeric antigen receptors (CARs) improve outcomes for CD19-expressing B cell malignancies. We evaluated a human application of T cells that were genetically modified using the Sleeping Beauty (SB) transposon/transposase system to express a CD19-specific CAR. METHODS: T cells were genetically modified using DNA plasmids from the SB platform to stably express a second-generation CD19-specific CAR and selectively propagated ex vivo with activating and propagating cells (AaPCs) and cytokines...
September 1, 2016: Journal of Clinical Investigation
Rui Liu, Ping Zhang, Yiqi Su, Huixing Lin, Hui Zhang, Lei Yu, Zhe Ma, Hongjie Fan
The mariner-based Himar1 system has been utilized for creating mutant libraries of many Gram-positive bacteria. Streptococcus suis serotype 2 (SS2) and Streptococcus equi ssp. zooepidemicus (SEZ) are primary pathogens of swine that threaten the swine industry in China. To provide a forward-genetics technology for finding virulent phenotype-related genes in these two pathogens, we constructed a novel temperature-sensitive suicide shuttle plasmid, pMar4s, which contains the Himar1 system transposon, TnYLB-1, and the Himar1 C9 transposase from pMarA and the repTAs temperature-sensitive fragment from pSET4s...
2016: Scientific Reports
Nigel P Minton, Muhammad Ehsaan, Christopher M Humphreys, Gareth T Little, Jonathan Baker, Anne M Henstra, Fungmin Liew, Michelle L Kelly, Lili Sheng, Katrin Schwarz, Ying Zhang
Clostridium species are both heroes and villains. Some cause serious human and animal diseases, those present in the gut microbiota generally contribute to health and wellbeing, while others represent useful industrial chassis for the production of chemicals and fuels. To understand, counter or exploit, there is a fundamental requirement for effective systems that may be used for directed or random genome modifications. We have formulated a simple roadmap whereby the necessary gene systems maybe developed and deployed...
May 24, 2016: Anaerobe
Elizabeth R Morris, Heather Grey, Grant McKenzie, Anita C Jones, Julia M Richardson
Cut-and-paste DNA transposons of the mariner/Tc1 family are useful tools for genome engineering and are inserted specifically at TA target sites. A crystal structure of the mariner transposase Mos1 (derived from Drosophila mauritiana), in complex with transposon ends covalently joined to target DNA, portrays the transposition machinery after DNA integration. It reveals severe distortion of target DNA and flipping of the target adenines into extra-helical positions. Fluorescence experiments confirm dynamic base flipping in solution...
2016: ELife
Antonio Palazzo, Domenica Lovero, Pietro D'Addabbo, Ruggiero Caizzi, René Massimiliano Marsano
Bari elements are members of the Tc1-mariner superfamily of DNA transposons, originally discovered in Drosophila melanogaster, and subsequently identified in silico in 11 sequenced Drosophila genomes and as experimentally isolated in four non-sequenced Drosophila species. Bari-like elements have been also studied for their mobility both in vivo and in vitro. We analyzed 23 Drosophila genomes and carried out a detailed characterization of the Bari elements identified, including those from the heterochromatic Bari1 cluster in D...
2016: PloS One
Anelis Maria Marin, Jésus de la Torre, Alfredo Ricardo Marques Oliveira, Andersson Barison, Leda Satie Chubatsu, Rose Adele Monteiro, Fabio de Oliveira Pedrosa, Emanuel Maltempi de Souza, Roseli Wassem, Estrella Duque, Juan Luis Ramos
In this study a random mutant library of Herbaspirillum seropedicae SmR1 was constructed by Tn5 insertion and a mutant incapable of utilizing naringenin as a carbon source was isolated. The Tn5 transposon was found to be inserted in the fdeE gene (Hsero_1007), which encodes a monooxygenase. Two other mutant strains in fdeC (Hsero_1005) and fdeG (Hsero_1009) genes coding for a dioxygenase and a putative cyclase, respectively, were obtained by site-directed mutagenesis and then characterized. LC-MS/MS analyses of culture supernatant from the fdeE mutant strain revealed that naringenin remained unaltered, suggesting that the FdeE protein is involved in the initial step of naringenin degradation...
April 5, 2016: Environmental Microbiology
Franka Voigt, Lisa Wiedemann, Cecilia Zuliani, Irma Querques, Attila Sebe, Lajos Mátés, Zsuzsanna Izsvák, Zoltán Ivics, Orsolya Barabas
Sleeping Beauty (SB) is a prominent Tc1/mariner superfamily DNA transposon that provides a popular genome engineering tool in a broad range of organisms. It is mobilized by a transposase enzyme that catalyses DNA cleavage and integration at short specific sequences at the transposon ends. To facilitate SB's applications, here we determine the crystal structure of the transposase catalytic domain and use it to model the SB transposase/transposon end/target DNA complex. Together with biochemical and cell-based transposition assays, our structure reveals mechanistic insights into SB transposition and rationalizes previous hyperactive transposase mutations...
March 30, 2016: Nature Communications
Ying Zhang, Shu Xu, Changsheng Chai, Sheng Yang, Weihong Jiang, Nigel P Minton, Yang Gu
Clostridium acetobutylicum is an industrially important Gram-positive organism, which is capable of producing economically important chemicals in the ABE (Acetone, Butanol and Ethanol) fermentation process. Renewed interests in the ABE process necessitate the availability of additional genetics tools to facilitate the derivation of a greater understanding of the underlying metabolic and regulatory control processes in operation through forward genetic strategies. In this study, a xylose inducible, mariner-based, transposon system was developed and shown to allow high-efficient random mutagenesis in the model strain ATCC 824...
April 2016: FEMS Microbiology Letters
Umberto Rosani, Alberto Pallavicini, Paola Venier
Small non-coding RNAs include powerful regulators of gene expression, transposon mobility and virus activity. Among the various categories, mature microRNAs (miRNAs) guide the translational repression and decay of several targeted mRNAs. The biogenesis of miRNAs depends on few gene products, essentially conserved from basal to higher metazoans, whose protein domains allow specific interactions with dsRNA. Here, we report the identification of key genes responsible of the miRNA biogenesis in 32 bivalves, with particular attention to the aquaculture species Mytilus galloprovincialis and Crassostrea gigas...
2016: PeerJ
Victoria Michael, Oliver Frank, Pascal Bartling, Carmen Scheuner, Markus Göker, Henner Brinkmann, Jörn Petersen
Alphaproteobacteria of the metabolically versatile Roseobacter group (Rhodobacteraceae) are abundant in marine ecosystems and represent dominant primary colonizers of submerged surfaces. Motility and attachment are the prerequisite for the characteristic 'swim-or-stick' lifestyle of many representatives such as Phaeobacter inhibens DSM 17395. It has recently been shown that plasmid curing of its 65-kb RepA-I-type replicon with >20 genes for exopolysaccharide biosynthesis including a rhamnose operon results in nearly complete loss of motility and biofilm formation...
October 2016: ISME Journal
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