Journal articles: 'Bacterial genetic transformation'

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HÅVARSTEIN, LEIV SIGVE. "Bacterial Gene Transfer by Natural Genetic Transformation." APMIS 106, S84 (November 1998): 43–46. http://dx.doi.org/10.1111/j.1600-0463.1998.tb05647.x.

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Draghi, Jeremy A., and Paul E. Turner. "DNA secretion and gene-level selection in bacteria." Microbiology 152, no. 9 (September 1, 2006): 2683–88. http://dx.doi.org/10.1099/mic.0.29013-0.

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Natural genetic transformation can facilitate gene transfer in many genera of bacteria and requires the presence of extracellular DNA. Although cell lysis can contribute to this extracellular DNA pool, several studies have suggested that the secretion of DNA from living bacteria may also provide genetic material for transformation. This paper reviews the evidence for specific secretion of DNA from intact bacteria into the extracellular environment and examines this behaviour from a population-genetics perspective. A mathematical model demonstrates that the joint action of DNA secretion and transformation creates a novel type of gene-level natural selection. This model demonstrates that gene-level selection could explain the existence of DNA secretion mechanisms that provide no benefit to individual cells or populations of bacteria. Additionally, the model predicts that any trait affecting DNA secretion will experience selection at the gene level in a transforming population. This analysis confirms that the secretion of DNA from intact bacterial cells is fully explicable with evolutionary theory, and reveals a novel mechanism for bacterial evolution.

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Engelstädter, Jan, and Danesh Moradigaravand. "Adaptation through genetic time travel? Fluctuating selection can drive the evolution of bacterial transformation." Proceedings of the Royal Society B: Biological Sciences 281, no. 1775 (January 22, 2014): 20132609. http://dx.doi.org/10.1098/rspb.2013.2609.

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Natural transformation is a process whereby bacteria actively take up DNA from the surrounding environment and incorporate it into their genome. Natural transformation is widespread in bacteria, but its evolutionary significance is still debated. Here, we hypothesize that transformation may confer a fitness advantage in changing environments through a process we term ‘genetic time travel’: by taking up old genes that were retained in the environment, the bacteria may revert to a past genotypic state that proves advantageous in the present or a future environment. We scrutinize our hypothesis by means of a mathematical model involving two bacterial types (transforming and non-transforming), a single locus under natural selection and a free DNA pool. The two bacterial types were competed in environments with changing selection regimes. We demonstrate that for a wide range of parameter values for the DNA turnover rate, the transformation rate and the frequency of environmental change, the transforming type outcompetes the non-transforming type. We discuss the empirical plausibility of our hypothesis, as well as its relationship to other hypotheses for the evolution of transformation in bacteria and sex more generally, speculating that ‘genetic time travel’ may also be relevant in eukaryotes that undergo horizontal gene transfer.

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Ambur, Ole Herman, Jan Engelstädter, Pål J. Johnsen, Eric L. Miller, and Daniel E. Rozen. "Steady at the wheel: conservative sex and the benefits of bacterial transformation." Philosophical Transactions of the Royal Society B: Biological Sciences 371, no. 1706 (October 19, 2016): 20150528. http://dx.doi.org/10.1098/rstb.2015.0528.

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Many bacteria are highly sexual, but the reasons for their promiscuity remain obscure. Did bacterial sex evolve to maximize diversity and facilitate adaptation in a changing world, or does it instead help to retain the bacterial functions that work right now? In other words, is bacterial sex innovative or conservative? Our aim in this review is to integrate experimental, bioinformatic and theoretical studies to critically evaluate these alternatives, with a main focus on natural genetic transformation, the bacterial equivalent of eukaryotic sexual reproduction. First, we provide a general overview of several hypotheses that have been put forward to explain the evolution of transformation. Next, we synthesize a large body of evidence highlighting the numerous passive and active barriers to transformation that have evolved to protect bacteria from foreign DNA, thereby increasing the likelihood that transformation takes place among clonemates. Our critical review of the existing literature provides support for the view that bacterial transformation is maintained as a means of genomic conservation that provides direct benefits to both individual bacterial cells and to transformable bacterial populations. We examine the generality of this view across bacteria and contrast this explanation with the different evolutionary roles proposed to maintain sex in eukaryotes. This article is part of the themed issue ‘Weird sex: the underappreciated diversity of sexual reproduction’.

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Udayabhanu, Jinu, Tiandai Huang, Shichao Xin, Jing Cheng, Yuwei Hua, and Huasun Huang. "Optimization of the Transformation Protocol for Increased Efficiency of Genetic Transformation in Hevea brasiliensis." Plants 11, no. 8 (April 14, 2022): 1067. http://dx.doi.org/10.3390/plants11081067.

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The recurring growth of bacterium in newly developed resistant cells and a minimal level of bacterial infection rate are the main limiting factors of Agrobacterium-mediated transformation experiments in Hevea brasiliensis. The current study aimed to optimize crucial factors of the transformation protocol in order to obtain an efficient transformation experimental model for Hevea using cotyledonary somatic embryos as explants. Transformation conditions such as antibiotic concentration, preculture duration, Agrobacterium concentration, sonication and cocultivation conditions were analyzed using the binary vector pCAMBIA2301. Transient transformation was confirmed by GUS histochemical staining. The best transformation efficiency was observed when the explants were not cultured on a preculture medium that contained acetosyringone at a level of 100 μM. The best results were obtained using a bacterial density of 0.45 at OD 600 nm, 50 s of sonication of explants in a bacterial liquid culture and a total incubation time of 18 min in the same bacterial suspension. Transmission electron microscopical analysis confirmed the impacts of sonication on bacterial infection efficiency. Cocultivation conditions of 22 °C and 84 h of darkness were optimal for the transfer of T-DNA. Agrobacterium was eliminated with 500 mg/L of timentin, and the selection of transformants was performed using 100 mg/L of kanamycin in the selection medium. The presence of transgene was confirmed in the resistant embryos by polymerase chain reaction (PCR). The improved method of genetic transformation established in the present study will be useful for the introduction of foreign genes of interest into the Hevea genome for the breeding of this economically important plant species in the future.

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Beard, C. B., S. L. O'Neill, P. Mason, L. Mandelco, C. R. Woese, R. B. Tesh, F. F. Richards, and S. Aksoy. "Genetic transformation and phylogeny of bacterial symbionts from tsetse." Insect Molecular Biology 1, no. 3 (February 1993): 123–31. http://dx.doi.org/10.1111/j.1365-2583.1993.tb00113.x.

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Majewski, Jacek, Piotr Zawadzki, Paul Pickerill, Frederick M. Cohan, and Christopher G. Dowson. "Barriers to Genetic Exchange between Bacterial Species: Streptococcus pneumoniae Transformation." Journal of Bacteriology 182, no. 4 (February 15, 2000): 1016–23. http://dx.doi.org/10.1128/jb.182.4.1016-1023.2000.

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ABSTRACT Interspecies genetic exchange is an important evolutionary mechanism in bacteria. It allows rapid acquisition of novel functions by transmission of adaptive genes between related species. However, the frequency of homologous recombination between bacterial species decreases sharply with the extent of DNA sequence divergence between the donor and the recipient. In Bacillus andEscherichia, this sexual isolation has been shown to be an exponential function of sequence divergence. Here we demonstrate that sexual isolation in transformation between Streptococcus pneumoniae recipient strains and donor DNA from related strains and species follows the described exponential relationship. We show that the Hex mismatch repair system poses a significant barrier to recombination over the entire range of sequence divergence (0.6 to 27%) investigated. Although mismatch repair becomes partially saturated, it is responsible for 34% of the observed sexual isolation. This is greater than the role of mismatch repair inBacillus but less than that in Escherichia. The remaining non-Hex-mediated barrier to recombination can be provided by a variety of mechanisms. We discuss the possible additional mechanisms of sexual isolation, in view of earlier findings fromBacillus, Escherichia, andStreptococcus.

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Lorenz, M. G., and W. Wackernagel. "Bacterial gene transfer by natural genetic transformation in the environment." Microbiological Reviews 58, no. 3 (1994): 563–602. http://dx.doi.org/10.1128/mmbr.58.3.563-602.1994.

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Lorenz, M. G., and W. Wackernagel. "Bacterial gene transfer by natural genetic transformation in the environment." Microbiological Reviews 58, no. 3 (1994): 563–602. http://dx.doi.org/10.1128/mr.58.3.563-602.1994.

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Johnston, Christopher D., Sean L. Cotton, Susan R. Rittling, Jacqueline R. Starr, Gary G. Borisy, Floyd E. Dewhirst, and Katherine P. Lemon. "Systematic evasion of the restriction-modification barrier in bacteria." Proceedings of the National Academy of Sciences 116, no. 23 (May 16, 2019): 11454–59. http://dx.doi.org/10.1073/pnas.1820256116.

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Bacteria that are recalcitrant to genetic manipulation using modern in vitro techniques are termed genetically intractable. Genetic intractability is a fundamental barrier to progress that hinders basic, synthetic, and translational microbiology research and development beyond a few model organisms. The most common underlying causes of genetic intractability are restriction-modification (RM) systems, ubiquitous defense mechanisms against xenogeneic DNA that hinder the use of genetic approaches in the vast majority of bacteria and exhibit strain-level variation. Here, we describe a systematic approach to overcome RM systems. Our approach was inspired by a simple hypothesis: if a synthetic piece of DNA lacks the highly specific target recognition motifs for a host’s RM systems, then it is invisible to these systems and will not be degraded during artificial transformation. Accordingly, in this process, we determine the genome and methylome of an individual bacterial strain and use this information to define the bacterium’s RM target motifs. We then synonymously eliminate RM targets from the nucleotide sequence of a genetic tool in silico, synthesize an RM-silent “SyngenicDNA” tool, and propagate the tool as minicircle plasmids, termed SyMPL (SyngenicDNA Minicircle Plasmid) tools, before transformation. In a proof-of-principle of our approach, we demonstrate a profound improvement (five orders of magnitude) in the transformation of a clinically relevant USA300 strain of Staphylococcus aureus. This stealth-by-engineering SyngenicDNA approach is effective, flexible, and we expect in future applications could enable microbial genetics free of the restraints of restriction-modification barriers.

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Stevenson, Brian, James L. Bono, Abdallah Elias, Kit Tilly, and Patricia Rosa. "Transformation of the Lyme Disease Spirochete Borrelia burgdorferi with Heterologous DNA." Journal of Bacteriology 180, no. 18 (September 15, 1998): 4850–55. http://dx.doi.org/10.1128/jb.180.18.4850-4855.1998.

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ABSTRACT Studies of the spirochete Borrelia burgdorferi have been hindered by the scarcity of genetic tools that can be used in these bacteria. For the first time, a method has been developed by which heterologous DNA (DNA without a naturally occurring B. burgdorferi homolog) can be introduced into and persistently maintained by B. burgdorferi. This technique uses integration of circular DNA into the bacterial genome via a single-crossover event. The ability to transform B. burgdorferi with heterologous DNA will now permit a wide range of experiments on the biology of these bacteria and their involvement in the many facets of Lyme disease.

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Hnatiuk, I. S., O. I. Varchenko, M. F. Parii, and Yu V. Symonenko. "Creation of a genetic vector carrying a synthesis bacterial protein gene CAS9 for plant genome editing." Faktori eksperimental'noi evolucii organizmiv 26 (September 1, 2020): 176–82. http://dx.doi.org/10.7124/feeo.v26.1263.

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Aim. To create a genetic construct carrying the bacterial protein Cas9 gene, the reporter β-glucuronidase gus gene, as well as the marker phosphinotricin-N-acetyltransferase bar gene for plant genome editing. Methods. Molecular-biological, biotechnological, microbiological and bioinformatic methods were used in the study; Golden Gate molecular cloning method was used to create genetic constructs. Results. The genetic construct pSPE2053 which carries the Cas9 endonuclease gene, the gus and bar genes was created; the assembly correctness of all vector elements was confirmed by polymerase chain reaction; the construct was transferred to Escherichia coli and Agrobacterium tumefaciens cells; β-glucuronidase gene expression was verified by histochemical analysis after Nicotiana rustica L transient genetic transformation. Conclusions. The created genetic construct can be used to edit the plant genome for both stable and transient genetic transformation to accumulate recombinant Cas9 protein. The guide RNA sequences may be subsequently transferred into such plants using either stable or transient genetic transformation or traditional crossing methods. Keywords: cloning, genetic construction, gus and bar genes, Cas9 endonuclease protein, transient expression.

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Lam, Trinh, Martin D. Brennan, Donald A. Morrison, and David T. Eddington. "Femtoliter droplet confinement ofStreptococcus pneumoniae: bacterial genetic transformation by cell–cell interaction in droplets." Lab on a Chip 19, no. 4 (2019): 682–92. http://dx.doi.org/10.1039/c8lc01367e.

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Brimacombe, Cedric A., Hao Ding, Jeanette A. Johnson, and J. Thomas Beatty. "Homologues of Genetic Transformation DNA Import Genes Are Required for Rhodobacter capsulatus Gene Transfer Agent Recipient Capability Regulated by the Response Regulator CtrA." Journal of Bacteriology 197, no. 16 (June 1, 2015): 2653–63. http://dx.doi.org/10.1128/jb.00332-15.

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ABSTRACTGene transfer agents (GTAs) morphologically resemble small, double-stranded DNA (dsDNA) bacteriophages; however, their only known role is to package and transfer random pieces of the producing cell genome to recipient cells. The best understood GTA is that ofRhodobacter capsulatus, termed RcGTA. We discovered that homologues of three genes involved in natural transformation in other bacteria,comEC,comF, andcomM, are essential for RcGTA-mediated gene acquisition. This paper gives genetic and biochemical evidence that RcGTA-borne DNA entry into cells requires the ComEC and ComF putative DNA transport proteins and genetic evidence that putative cytoplasmic ComM protein of unknown function is required for recipient capability. Furthermore, the master regulator of RcGTA production in <1% of a cell population, CtrA, which is also required for gene acquisition in recipient cells, is expressed in the vast majority of the population. Our results indicate that RcGTA-mediated gene transfer combines key aspects of two bacterial horizontal gene transfer mechanisms, where donor DNA is packaged in transducing phage-like particles and recipient cells take up DNA using natural transformation-related machinery. Both of these differentiated subsets of a culture population, donors and recipients, are dependent on the same response regulator, CtrA.IMPORTANCEHorizontal gene transfer (HGT) is a major driver of bacterial evolution and adaptation to environmental stresses. Traits such as antibiotic resistance or metabolic properties can be transferred between bacteria via HGT; thus, HGT can have a tremendous effect on the fitness of a bacterial population. The three classically described HGT mechanisms are conjugation, transformation, and phage-mediated transduction. More recently, the HGT factor GTA was described, where random pieces of producing cell genome are packaged into phage-like particles that deliver DNA to recipient cells. In this report, we show that transport of DNA borne by theR. capsulatusRcGTA into recipient cells requires key genes previously thought to be specific to natural transformation pathways. These findings indicate that RcGTA combines central aspects of phage-mediated transduction and natural transformation in an efficient, regulated mode of HGT.

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Finkel, Steven E., and Roberto Kolter. "DNA as a Nutrient: Novel Role for Bacterial Competence Gene Homologs." Journal of Bacteriology 183, no. 21 (November 1, 2001): 6288–93. http://dx.doi.org/10.1128/jb.183.21.6288-6293.2001.

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ABSTRACT The uptake and stable maintenance of extracellular DNA, genetic transformation, is universally recognized as a major force in microbial evolution. We show here that extracellular DNA, both homospecific and heterospecific, can also serve as the sole source of carbon and energy supporting microbial growth. Mutants unable to consume DNA suffer a significant loss of fitness during stationary-phase competition. InEscherichia coli, the use of DNA as a nutrient depends on homologs of proteins involved in natural genetic competence and transformation in Haemophilus influenzae andNeisseria gonorrhoeae. Homologs of these E. coli genes are present in many members of the γ subclass ofProteobacteria, suggesting that the mechanisms for consumption of DNA may have been widely conserved during evolution.

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Lacroix, Benoît, and Vitaly Citovsky. "Pathways of DNA Transfer to Plants fromAgrobacterium tumefaciensand Related Bacterial Species." Annual Review of Phytopathology 57, no. 1 (August 25, 2019): 231–51. http://dx.doi.org/10.1146/annurev-phyto-082718-100101.

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Genetic transformation of host plants by Agrobacterium tumefaciens and related species represents a unique model for natural horizontal gene transfer. Almost five decades of studying the molecular interactions between Agrobacterium and its host cells have yielded countless fundamental insights into bacterial and plant biology, even though several steps of the DNA transfer process remain poorly understood. Agrobacterium spp. may utilize different pathways for transferring DNA, which likely reflects the very wide host range of Agrobacterium. Furthermore, closely related bacterial species, such as rhizobia, are able to transfer DNA to host plant cells when they are provided with Agrobacterium DNA transfer machinery and T-DNA. Homologs of Agrobacterium virulence genes are found in many bacterial genomes, but only one non- Agrobacterium bacterial strain, Rhizobium etli CFN42, harbors a complete set of virulence genes and can mediate plant genetic transformation when carrying a T-DNA-containing plasmid.

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Deutch, Charles E. "Transformation of Escherichia coli with the pGLO Plasmid: Going beyond the Kit." American Biology Teacher 81, no. 1 (January 1, 2019): 52–55. http://dx.doi.org/10.1525/abt.2019.81.1.52.

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The Bio-Rad pGLO bacterial transformation kit is commonly used to demonstrate this form of genetic exchange, which occurs in bacteria and eukaryotes and which differs fundamentally from transduction and conjugation. The basic experiment leads to the formation of green fluorescent colonies of Escherichia coli and can be extended to illustrate the specificity of the interaction between sugars and the AraC protein, the phenomenon of carbon catabolite repression, the substrate specificity of the β-lactamase encoded by the plasmid, and the role of host restriction/modification systems in the transformation process. pGLO DNA also can be isolated using plasmid mini-prep kits, analyzed with restriction endonucleases, and used to study the conditions for transformation in more detail.

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Satish, Lakkakula, Madhu Kamle, Guy Keren, Chandrashekhar D. Patil, Galit Yehezkel, Ze’ev Barak, Varda Kagan-Zur, Ariel Kushmaro, and Yaron Sitrit. "Agrobacterium tumefaciens-Mediated Genetic Transformation of the Ect-endomycorrhizal Fungus Terfezia boudieri." Genes 11, no. 11 (October 30, 2020): 1293. http://dx.doi.org/10.3390/genes11111293.

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Mycorrhizal desert truffles such as Terfezia boudieri, Tirmania nivea, and Terfezia claveryi, form mycorrhizal associations with plants of the Cistaceae family. These valued truffles are still collected from the wild and not cultivated under intensive farming due to the lack of basic knowledge about their biology at all levels. Recently, several genomes of desert truffles have been decoded, enabling researchers to attempt genetic manipulations to enable cultivation. To execute such manipulations, the development of molecular tools for genes transformation into truffles is needed. We developed an Agrobacterium tumefaciens-mediated genetic transformation system in T. boudieri. This system was optimized for the developmental stage of the mycelia explants, bacterial optical density, infection and co-cultivation durations, and concentrations of the selection antibiotics. The pFPL-Rh plasmid harboring hph gene conferring hygromycin resistance as a selection marker and the red fluorescent protein gene were used as visual reporters. The optimal conditions were incubation with 200 μM of acetosyringone, attaining a bacterial optical density of 0.3 OD600; transfer time of 45 min; and co-cultivation for 3 days. This is the first report on a transformation system for T. boudieri, and the proposed protocol can be adapted for the transformation of other important desert truffles as well as ectomycorrhizal species.

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Gallagher, Larry A., Jeannie Bailey, and Colin Manoil. "Ranking essential bacterial processes by speed of mutant death." Proceedings of the National Academy of Sciences 117, no. 30 (July 14, 2020): 18010–17. http://dx.doi.org/10.1073/pnas.2001507117.

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Mutant phenotype analysis of bacteria has been revolutionized by genome-scale screening procedures, but essential genes have been left out of such studies because mutants are missing from the libraries analyzed. Since essential genes control the most fundamental processes of bacterial life, this is a glaring deficiency. To address this limitation, we developed a procedure for transposon insertion mutant sequencing that includes essential genes. The method, called transformation transposon insertion mutant sequencing (TFNseq), employs saturation-level libraries of bacterial mutants generated by natural transformation with chromosomal DNA mutagenized heavily by in vitro transposition. The efficient mutagenesis makes it possible to detect large numbers of insertions in essential genes immediately after transformation and to follow their loss during subsequent growth. It was possible to order 45 essential processes based on how rapidly their inactivation inhibited growth. Inactivating ATP production, deoxyribonucleotide synthesis, or ribosome production blocked growth the fastest, whereas inactivating cell division or outer membrane protein synthesis blocked it the slowest. Individual mutants deleted of essential loci formed microcolonies of nongrowing cells whose sizes were generally consistent with the TFNseq ordering. The sensitivity of essential functions to genetic inactivation provides a metric for ranking their relative importance for bacterial replication and growth. Highly sensitive functions could represent attractive antibiotic targets since even partial inhibition should reduce growth.

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Dandekar, Abhaya M., Lori A. Martin, and Gale McGranahan. "Genetic Transformation and Foreign Gene Expression in Walnut Tissue." Journal of the American Society for Horticultural Science 113, no. 6 (November 1988): 945–49. http://dx.doi.org/10.21273/jashs.113.6.945.

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Abstract A system was developed to transform walnut cultivars using the natural gene transfer system of Agrobacterium tumefaciens. We report the infection of English walnut (Juglans regia L.), northern California black walnut (Juglans hindsii), and their F1 hybrid ‘Paradox’ with A. tumefaciens carrying various recombinant derivatives of the tumor-inducing (Ti) plasmids, pTiA6 and pTiB6S3. The three walnut species, each represented by a single micropropagated clone, were found to be equally susceptible to Agrobacterium-induced tumor formation in vitro. Stable lines were established from tumors induced on each clone, and, unlike normal stem callus, these tumor cells grew rapidly in culture media without exogenous plant hormones. High-voltage paper electrophoretic analysis revealed the presence of opines in the walnut tumor tissue. The presence of a foreign gene was demonstrated by expression of a chimeric bacterial gene that encodes resistance to the antibiotic kanamycin, and also by the presence of foreign DNA sequences in genomic DNA isolated from tumors.

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Lacroix, Benot, and Vitaly Citovsky. "The roles of bacterial and host plant factors in Agrobacterium-mediated genetic transformation." International Journal of Developmental Biology 57, no. 6-7-8 (2013): 467–81. http://dx.doi.org/10.1387/ijdb.130199bl.

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Tanaka, Kohjiro, Seiichi Furukawa, Naruo Nikoh, Tetsuhiko Sasaki, and Takema Fukatsu. "Complete WO Phage Sequences Reveal Their Dynamic Evolutionary Trajectories and Putative Functional Elements Required for Integration into the Wolbachia Genome." Applied and Environmental Microbiology 75, no. 17 (July 10, 2009): 5676–86. http://dx.doi.org/10.1128/aem.01172-09.

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ABSTRACT Wolbachia endosymbionts are ubiquitously found in diverse insects including many medical and hygienic pests, causing a variety of reproductive phenotypes, such as cytoplasmic incompatibility, and thereby efficiently spreading in host insect populations. Recently, Wolbachia-mediated approaches to pest control and management have been proposed, but the application of these approaches has been hindered by the lack of genetic transformation techniques for symbiotic bacteria. Here, we report the genome and structure of active bacteriophages from a Wolbachia endosymbiont. From the Wolbachia strain wCauB infecting the moth Ephestia kuehniella two closely related WO prophages, WOcauB2 of 43,016 bp with 47 open reading frames (ORFs) and WOcauB3 of 45,078 bp with 46 ORFs, were characterized. In each of the prophage genomes, an integrase gene and an attachment site core sequence were identified, which are putatively involved in integration and excision of the mobile genetic elements. The 3′ region of the prophages encoded genes with sequence motifs related to bacterial virulence and protein-protein interactions, which might represent effector molecules that affect cellular processes and functions of their host bacterium and/or insect. Database searches and phylogenetic analyses revealed that the prophage genes have experienced dynamic evolutionary trajectories. Genes similar to the prophage genes were found across divergent bacterial phyla, highlighting the active and mobile nature of the genetic elements. We suggest that the active WO prophage genomes and their constituent sequence elements would provide a clue to development of a genetic transformation vector for Wolbachia endosymbionts.

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Bello-López, J. Manuel, Omar A. Cabrero-Martínez, Gabriela Ibáñez-Cervantes, Cecilia Hernández-Cortez, Leda I. Pelcastre-Rodríguez, Luis U. Gonzalez-Avila, and Graciela Castro-Escarpulli. "Horizontal Gene Transfer and Its Association with Antibiotic Resistance in the Genus Aeromonas spp." Microorganisms 7, no. 9 (September 18, 2019): 363. http://dx.doi.org/10.3390/microorganisms7090363.

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The evolution of multidrug resistant bacteria to the most diverse antimicrobials known so far pose a serious problem to global public health. Currently, microorganisms that develop resistant phenotypes to multiple drugs are associated with high morbidity and mortality. This resistance is encoded by a group of genes termed ‘bacterial resistome’, divided in intrinsic and extrinsic resistome. The first one refers to the resistance displayed on an organism without previous exposure to an antibiotic not involving horizontal genetic transfer, and it can be acquired via mutations. The latter, on the contrary, is acquired exclusively via horizontal genetic transfer involving mobile genetic elements that constitute the ‘bacterial mobilome’. This transfer is mediated by three different mechanisms: transduction, transformation, and conjugation. Recently, a problem of public health due to implications in the emergence of multi-drug resistance in Aeromonas spp. strains in water environments has been described. This is derived from the genetic material transfer via conjugation events. This is important, since bacteria that have acquired antibiotic resistance in natural environments can cause infections derived from their ingestion or direct contact with open wounds or mucosal tissue, which in turn, by their resistant nature, makes their eradication complex. Implications of the emergence of resistance in Aeromonas spp. by horizontal gene transfer on public health are discussed.

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Kung, Stephanie H., and Rodrigo P. P. Almeida. "Natural Competence and Recombination in the Plant Pathogen Xylella fastidiosa." Applied and Environmental Microbiology 77, no. 15 (June 10, 2011): 5278–84. http://dx.doi.org/10.1128/aem.00730-11.

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ABSTRACTHomologous recombination is one of many forces contributing to the diversity, adaptation, and emergence of pathogens. For naturally competent bacteria, transformation is one possible route for the acquisition of novel genetic material. This study demonstrates thatXylella fastidiosa, a generalist bacterial plant pathogen responsible for many emerging plant diseases, is naturally competent and able to homologously recombine exogenous DNA into its genome. Several factors that affect transformation and recombination efficiencies, such as nutrient availability, growth stage, and methylation of transforming DNA, were identified. Recombination was observed in at least one out of every 106cells when exogenous plasmid DNA was supplied and one out of every 107cells when different strains were grown togetherin vitro. Based on previous genomic studies and experimental data presented here, there is mounting evidence that recombination can occur at relatively high rates and could play a large role in shaping the genetic diversity ofX. fastidiosa.

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Kung, Stephanie H., and Rodrigo P. P. Almeida. "Biological and genetic factors regulating natural competence in a bacterial plant pathogen." Microbiology 160, no. 1 (January 1, 2014): 37–46. http://dx.doi.org/10.1099/mic.0.070581-0.

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For naturally competent bacteria, spatially structured growth can provide an environment for enhanced horizontal gene transfer through transformation and recombination. DNA is often present in the extracellular environment, such as in the extracellular matrix of biofilms, and the lysis of a single cell can result in high local DNA concentrations. Xylella fastidiosa is a naturally competent plant pathogen that typically lives in a surface-attached state, yet previous work characterizing the competence of this organism was conducted with planktonic cells in liquid environments. Here, we show that transformation and recombination efficiencies are two to three orders of magnitude higher for cells grown on solid compared with liquid media, with maximum recombination efficiencies of about 10−3. Cells were highly competent throughout their exponential growth phase, with no significant change in recombination efficiencies until population growth rates began to slow. Mutations in type IV pili, competency-related, and cell–cell signalling genes significantly impacted the ability of X. fastidiosa to acquire and incorporate DNA. Because X. fastidiosa is highly competent when growing in a surface-attached state, as it does within its insect vectors and host plants, recombination of naturally transformed DNA could be a significant route by which horizontal gene transfer occurs in natural environments.

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Glick, Baernard R., Heather E. Brooks, and J. J. Pasternak. "Physiological effects of plasmid DNA transformation on Azotobacter vinelandii." Canadian Journal of Microbiology 32, no. 2 (February 1, 1986): 145–48. http://dx.doi.org/10.1139/m86-028.

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Genetic transformation of Azotobacter vinelandii by the introduction of broad-host-range plasmid DNA (i.e., pRK2501, RSF1010, or pGSS15) causes a number of physiological changes. As shown here, the capacity for nitrogen fixation, mean cell size, and synthesis of siderophores are decreased, whereas the production of capsular slime is enhanced. These findings suggest that the presence of plasmid DNA imposes a "metabolic load" on Azotobacter vinelandii. Therefore, it cannot be assumed a priori that the introduction of plasmid DNA into Azotobacter vinelandii will not disrupt some normal physiological processes. The implications of these findings are discussed, specifically in the context of developing Azotobacter vinelandii as an effective bacterial fertilizer by genetic manipulation.

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García, Belén, Elías R. Olivera, Ángel Sandoval, Elsa Arias-Barrau, Sagrario Arias, Germán Naharro, and José M. Luengo. "Strategy for Cloning Large Gene Assemblages as Illustrated Using the Phenylacetate and Polyhydroxyalkanoate Gene Clusters." Applied and Environmental Microbiology 70, no. 8 (August 2004): 5019–25. http://dx.doi.org/10.1128/aem.70.8.5019-5025.2004.

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ABSTRACT We report an easy procedure for isolating chromosome-clustered genes. By following this methodology, the entire set of genes belonging to the phenylacetic acid (PhAc; 18-kb) pathway as well as those required for the synthesis and mobilization of different polyhydroxyalkanoates (PHAs; 6.4 kb) in Pseudomonas putida U were recovered directly from the bacterial chromosome and cloned into a plasmid for the first time. The transformation of different bacteria with these genetic constructions conferred on them the ability to either degrade PhAc or synthesize bioplastics (PHAs).

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Cousins, YL, BR Lyon, and DJ Llewellyn. "Transformation of an Australian Cotton Cultivar: Prospects for Cotton Improvement Through Genetic Engineering." Functional Plant Biology 18, no. 5 (1991): 481. http://dx.doi.org/10.1071/pp9910481.

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Somatic embryogenesis and regeneration of whole plants is a highly genotype-dependent process in cotton. We have identified at least one highly regenerable Australian cultivar, Siokra 1-3, which is a sister line to the current major variety being grown in Australia. A number of plants have been regenerated and although some are showing abnormal pollen development, most can produce fertile seed when selfed or crossed with a normal pollen donor. Agrobacterium tumefaciens has been used to efficiently produce fertile transgenic Siokra 1-3 plants expressing novel genes such as the bacterial neomycin phosphotransferase or the β-glucuronidase. This is the first example of the transformation of an elite commercial cultivar. Critical factors in the transformation are the use of a supervirulent disarmed Ti-plasmid with a binary transformation vector, and a highly regenerable genotype of cotton. Bacterial concentration at the time of infection, tissue age, kanamycin selection regime, and co-cultivation support and media composition all have an influence on transformation efficiency and were optimised in our protocol. The ability to transform an elite Australian cultivar of cotton paves the way for agronomic improvements through genetic engineering. We have concentrated on increasing the tolerance of Australian cotton to the herbicide 2,4-D (to protect it from spray drift damage from adjacent cereal crops), and increasing its tolerance to insect pests, such as Helicoverpa armigera, using BT-toxin genes, protease inhibitors and other novel insect resistance genes.

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Salzer, Ralf, Friederike Joos, and Beate Averhoff. "Type IV Pilus Biogenesis, Twitching Motility, and DNA Uptake in Thermus thermophilus: Discrete Roles of Antagonistic ATPases PilF, PilT1, and PilT2." Applied and Environmental Microbiology 80, no. 2 (November 8, 2013): 644–52. http://dx.doi.org/10.1128/aem.03218-13.

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ABSTRACTNatural transformation has a large impact on lateral gene flow and has contributed significantly to the ecological diversification and adaptation of bacterial species.Thermus thermophilusHB27 has emerged as the leading model organism for studies of DNA transporters in thermophilic bacteria. Recently, we identified a zinc-binding polymerization nucleoside triphosphatase (NTPase), PilF, which is essential for the transport of DNA through the outer membrane. Here, we present genetic evidence that PilF is also essential for the biogenesis of pili. One of the most challenging questions was whetherT. thermophilushas any depolymerization NTPase acting as a counterplayer of PilF. We identified two depolymerization NTPases, PilT1 (TTC1621) and PilT2 (TTC1415), both of which are required for type IV pilus (T4P)-mediated twitching motility and adhesion but dispensable for natural transformation. This suggests that T4P dynamics are not required for natural transformation. The latter finding is consistent with our suggestion that inT. thermophilus, T4P and natural transformation are linked but distinct systems.

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Akter, Shirin, Sanjida Rahman Mollika, RH Sarker, and M. Imdadul Hoque. "Agrobacterium?mediated Genetic Transformation of Two Varieties of Brassica juncea (L.) Using Marker Genes." Plant Tissue Culture and Biotechnology 26, no. 1 (September 27, 2016): 55–65. http://dx.doi.org/10.3329/ptcb.v26i1.29767.

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Protocol for Agrobacterium?mediated genetic transformation using hypocotyl and cotyledonary leaf with petiole from two local varieties of Brassica juncea was established by optimizing various factors influencing transformation. GUS histochemical assay revealed that the cotyledonary leaf with petiole and hypocotyl explants had positive interaction with the Agrobacterium strain LBA4404 containing the binary plasmid pBI121 which has marker genes like, GUS and nptII. Maximum transformation was obtained with bacterial suspension having an optical density of 0.8 at 600 nm, 30 min of incubation and 72 hours of co?cultivation. The transient and stable integration of the marker genes were confirmed through histochemical GUS assay, as well as PCR analysis.Plant Tissue Cult. & Biotech. 26(1): 55-65, 2016 (June)

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Gong, Lisha, Min Lu, and Huaming An. "Generation of Composite Rosa roxburghii Plants with Transgenic Roots by Agrobacterium-Mediated Transformation." Horticulturae 8, no. 11 (November 16, 2022): 1079. http://dx.doi.org/10.3390/horticulturae8111079.

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Rosa roxburghii Tratt. is an emerging fruit endemic to China, which has the reputation of being the “King of Vitamin C” because of its abundance of vitamin C. However, it is also a recalcitrant species that imposes severe limitations on the transformation and whole-plant regeneration processes, restricting the verification of the functional genes. Therefore, developing a feasible and efficient genetic transformation method for R. roxburghii is an urgent requirement. Herein, K599 with eGFP was used as the Agrobacterium strain to optimize the genetic transformation from four factors: bacterial concentration, seedling age, infection site, and method. First, the original roots of 5-day-old seedlings were excised, and then the slant cuts of the remaining hypocotyls with 0.5 cm length were placed in K599 at an OD600 of 0.4. Subsequently, the explants were planted in a moistened sterile vermiculite after the beveled site was stained with a clump of bacteria. The results showed that the transformation efficiency of this cutting method was almost 28% at 30 days post-inoculation, while the transformation efficiency obtained by injecting 5-day-old seedlings 0.5–1.0 cm away from the primary root with K599 at an OD600 of 0.4 was only about 7%. Taken together, the current findings provide evidence that Agrobacterium-mediated transformation is a simple, fast, and efficient approach for generating composite R. roxburghii plants. Thus, this method has a broad application to analyze the gene functions in R. roxburghii and other related plant species.

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Bhattacharya, Arpita, Caroline Köhrer, Debabrata Mandal, and Uttam L. RajBhandary. "Nonsense suppression in archaea." Proceedings of the National Academy of Sciences 112, no. 19 (April 27, 2015): 6015–20. http://dx.doi.org/10.1073/pnas.1501558112.

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Bacterial strains carrying nonsense suppressor tRNA genes played a crucial role in early work on bacterial and bacterial viral genetics. In eukaryotes as well, suppressor tRNAs have played important roles in the genetic analysis of yeast and worms. Surprisingly, little is known about genetic suppression in archaea, and there has been no characterization of suppressor tRNAs or identification of nonsense mutations in any of the archaeal genes. Here, we show, using the β-gal gene as a reporter, that amber, ochre, and opal suppressors derived from the serine and tyrosine tRNAs of the archaeonHaloferax volcaniiare active in suppression of their corresponding stop codons. Using a promoter for tRNA expression regulated by tryptophan, we also show inducible and regulatable suppression of all three stop codons inH. volcanii. Additionally, transformation of aΔpyrE2 H. volcaniistrain with plasmids carrying the genes for apyrE2amber mutant and the serine amber suppressor tRNA yielded transformants that grow on agar plates lacking uracil. Thus, an auxotrophic amber mutation in thepyrE2gene can be complemented by expression of the amber suppressor tRNA. These results pave the way for generating archaeal strains carrying inducible suppressor tRNA genes on the chromosome and their use in archaeal and archaeviral genetics. We also provide possible explanations for why suppressor tRNAs have not been identified in archaea.

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Pasmawati, Pasmawati, Aris Tjahjoleksono, and Suharsono Suharsono. "Obtaining of transgenic potato (Solanum tuberosum L.) cultivar IPB CP3 containing LYZ‐C gene resistant to bacterial wilt disease." Indonesian Journal of Biotechnology 26, no. 1 (March 30, 2021): 48. http://dx.doi.org/10.22146/ijbiotech.61682.

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Bacterial wilt caused by Ralstonia solanacearum is one of the most important bacterial diseases in potato production. This study aimed to obtain the transgenic potato (Solanum tuberosum L.) cultivar IPB CP3, containing LYZ‐C gene encoding for lysozyme type C, resistant to bacterial disease caused by R. solanacearum. Genetic transformation using Agrobacterium tumefaciens LBA4404 to 124 internode explants resulted in the transformation efficiency of about 47.58% with a regeneration efficiency of approximately 30.51%. Gene integration analysis showed that 16 clones were confirmed as transgenic clones containing the LYZ‐C gene. Analysis of resistance to R. solanacearum of three transgenic clones showed that all three transgenic clones were more resistant than a non‐transgenic one. This result showed that the LYZ‐C gene integrated in the genome of transgenic potato increased the resistance of potato plants to R. solanacearum. We obtained two transgenic clones considered resistant to bacterial wilt disease.

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Hrahsel, Lalremsiami, Adreeja Basu, Lingaraj Sahoo, and Robert Thangjam. "Genetic Transformation of Musa acuminata cv. Vaibalhla (AAA) using Agrobacterium tumefaciens." Science & Technology Journal 5, no. 2 (July 1, 2017): 120–31. http://dx.doi.org/10.22232/stj.2017.05.02.09.

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Agrobacterium-mediated genetic transformation of Musa acuminata cv. Vaibalhla (AAA) was performed using Agrobacterium tumefaciens strain EHA105 harboring pCAMBIA2301AtNHX1 plasmid. For the transformation efficiency assay, the un-and pre-cultured immature male flower explants were subjected to various methods of injury such as hypodermal needle injury, with and without sonication and vacuum infiltration. The explants were then inoculated in bacterial suspension by occasional shaking for 30 minutes. After inoculation, explants were co-cultivated for 3 days in dark condition at 22° C in a liquid MS basal medium supplemented with 100 µM acetosyringone. For selection of transformants, the treated explants were cultured on Murashige and Skoog (MS) medium supplemented with BAP (2 mg/L), NAA (0.5 mg/L) and ascorbic acid (75 mg/L) along with antibiotics kanamycin (100 mg/L), cefotaxime (300 mg/L) and augmentin (300 mg/L).The putative transformation was analyzed using histochemical GUS assay. 14 and 21 days pre-cultured male flower explants subjected to 4-5 needle point injury resulted in 93.33% and 100% putative transformation respectively. 30s sonication combined with 5 minutes of vacuum infiltration for 7, 14 and 21 days pre-cultured immature male flower explants gave 73.33%, 66.66% and 71.42% putative transformation respectively.

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G., Anuradha, Mahendranath Gandikota, Krishnakanth Yadav T., Jagadish P., Raghurami Reddy M., Balachandran S.M.,, Siddiq E.A., and Yamini K.N. "Standardization of Agrobacterium mediated genetic transformation in Indica rice cv BPT-5204." Annals of Plant Sciences 7, no. 2 (January 31, 2018): 2037. http://dx.doi.org/10.21746/aps.2018.7.2.9.

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The BPT-5204 genotype from Indica rice cultivar (cv) is recalcitrant and showed low transformation frequency compare to japonica rice cv. Here we have optimized the efficient transformation protocol to minimize the time scale and enhance the transformation frequency by altering the key parameters like, acetosyringone (AS) concentration, optical density of bacterial culture and co-cultivation time. Highly proliferated 21 days old Scutellum derived embryogenic calli were infected with Agrobacterium tumefaciens harbouring pCAMBIA2300-Ds-En-Bar binary vector. T-DNA contains tetrameric CaMV35S enhancer elements along with bar gene which embedded between the transposable Ds element. At 0.5 OD600nm of Agrobacterium culture and co-cultivation for 2 days on MS co-cultivation medium containing 100 μM acetosyringone proved to be optimal and attained 6.2 % of transformation efficiency. The transformed calli and regenerated plantlets were proliferated on Murashige and Skoog (MS) medium containing phosphinothricin (PPT). Polymerase chain reaction (PCR) confirmed that intact T-DNA was successfully integrated in the rice genome. This protocol can be employed to develop transgenic rice plants with gain of functional mutagenesis.

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Gelvin, Stanton B. "Agrobacterium-Mediated Plant Transformation: the Biology behind the “Gene-Jockeying” Tool." Microbiology and Molecular Biology Reviews 67, no. 1 (March 2003): 16–37. http://dx.doi.org/10.1128/mmbr.67.1.16-37.2003.

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SUMMARY Agrobacterium tumefaciens and related Agrobacterium species have been known as plant pathogens since the beginning of the 20th century. However, only in the past two decades has the ability of Agrobacterium to transfer DNA to plant cells been harnessed for the purposes of plant genetic engineering. Since the initial reports in the early 1980s using Agrobacterium to generate transgenic plants, scientists have attempted to improve this “natural genetic engineer” for biotechnology purposes. Some of these modifications have resulted in extending the host range of the bacterium to economically important crop species. However, in most instances, major improvements involved alterations in plant tissue culture transformation and regeneration conditions rather than manipulation of bacterial or host genes. Agrobacterium-mediated plant transformation is a highly complex and evolved process involving genetic determinants of both the bacterium and the host plant cell. In this article, I review some of the basic biology concerned with Agrobacterium-mediated genetic transformation. Knowledge of fundamental biological principles embracing both the host and the pathogen have been and will continue to be key to extending the utility of Agrobacterium for genetic engineering purposes.

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Ando, Takafumi, Dawn A. Israel, Kazuo Kusugami, and Martin J. Blaser. "HP0333, a Member of the dprA Family, Is Involved in Natural Transformation in Helicobacter pylori." Journal of Bacteriology 181, no. 18 (September 15, 1999): 5572–80. http://dx.doi.org/10.1128/jb.181.18.5572-5580.1999.

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ABSTRACT Helicobacter pylori is naturally competent for DNA transformation, but the mechanism by which transformation occurs is not known. For Haemophilus influenzae, dprA is required for transformation by chromosomal but not plasmid DNA, and the complete genomic sequence of H. pylori 26695 revealed adprA homolog (HP0333). Examination of genetic databases indicates that DprA homologs are present in a wide variety of bacterial species. To examine whether HP0333 has a function similar todprA of H. influenzae, HP0333, present in each of 11 strains studied, was disrupted in two H. pyloriisolates. For both mutants, the frequency of transformation by H. pylori chromosomal DNA was markedly reduced, but not eliminated, compared to their wild-type parental strains. Mutation of HP0333 also resulted in a marked decrease in transformation frequency by a shuttle plasmid (pHP1), which differs from the phenotype described in H. influenzae. Complementation of the mutant with HP0333 inserted intrans in the chromosomal ureAB locus completely restored the frequency of transformation to that of the wild-type strain. Thus, while dprA is required for high-frequency transformation, transformation also may occur independently of DprA. The presence of DprA homologs in bacteria known not to be naturally competent suggests a broad function in DNA processing.

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Karnholz, Arno, Claudia Hoefler, Stefan Odenbreit, Wolfgang Fischer, Dirk Hofreuter, and Rainer Haas. "Functional and Topological Characterization of Novel Components of the comB DNA Transformation Competence System in Helicobacter pylori." Journal of Bacteriology 188, no. 3 (February 1, 2006): 882–93. http://dx.doi.org/10.1128/jb.188.3.882-893.2006.

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ABSTRACT Helicobacter pylori is one of the most diverse bacterial species known. A rational basis for this genetic variation may be provided by its natural competence for genetic transformation and high-frequency recombination. Many bacterial competence systems have homology with proteins that are involved in the assembly of type IV pili and type II secretion systems. In H. pylori, DNA uptake relies on a transport system related to type IV secretion systems (T4SS) designated the comB system. The prototype of a T4SS in Agrobacterium tumefaciens consists of 11 VirB proteins and VirD4, which form the core unit necessary for the delivery of single proteins or large nucleoprotein complexes into target cells. In the past we identified proteins ComB4 and ComB7 through ComB10 as being involved in the process of DNA uptake in H. pylori. In this study we identified and functionally characterized further (T4SS-homologous) components of the comB transformation competence system. By combining computer prediction modeling, experimental topology determination, generation of knockout strains, and genetic complementation studies we identified ComB2, ComB3, and ComB6 as essential components of the transformation apparatus, structurally and functionally homologous to VirB2, VirB3, and VirB6, respectively. comB2, comB3, and comB4 are organized as a separate operon. Thus, for the H. pylori comB system, all T4SS core components have been identified except for homologues to VirB1, VirD4, VirB5, and VirB11.

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Anand, Ajith, Zarir Vaghchhipawala, Choong-Min Ryu, Li Kang, Keri Wang, Olga del-Pozo, Gregory B. Martin, and Kirankumar S. Mysore. "Identification and Characterization of Plant Genes Involved in Agrobacterium-Mediated Plant Transformation by Virus-Induced Gene Silencing." Molecular Plant-Microbe Interactions® 20, no. 1 (January 2007): 41–52. http://dx.doi.org/10.1094/mpmi-20-0041.

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Genetic transformation of plant cells by Agrobacterium tu-mefaciens represents a unique case of trans-kingdom sex requiring the involvement of both bacterial virulence proteins and plant-encoded proteins. We have developed in planta and leaf-disk assays in Nicotiana benthamiana for identifying plant genes involved in Agrobacterium-mediated plant transformation using virus-induced gene silencing (VIGS) as a genomics tool. VIGS was used to validate the role of several genes that are either known or speculated to be involved in Agrobacterium-mediated plant transformation. We showed the involvement of a nodulin-like protein and an alpha-expansin protein (α-Exp) during Agrobacterium infection. Our data suggest that α-Exp is involved during early events of Agrobacterium-mediated transformation but not required for attaching A. tumefaciens. By employing the combination of the VIGS-mediated forward genetics approach and an in planta tumorigenesis assay, we identified 21 ACG (altered crown gall) genes that, when silenced, produced altered crown gall phenotypes upon infection with a tumorigenic strain of A. tumefaciens. One of the plant genes identified from the screening, Histone H3 (H3), was further characterized for its biological role in Agrobacterium-mediated plant transformation. We provide evidence for the role of H3 in transfer DNA integration. The data presented here suggest that the VIGS-based approach to identify and characterize plant genes involved in genetic transformation of plant cells by A. tumefaciens is simple, rapid, and robust and complements other currently used approaches.

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Qiu-Hua, Li, Hong Bo, Tong Zheng, Ma Chao, Guan Ai-Nong, Yu Jing-Juan, and Gao Jun-Ping. "Establishment of regeneration system and transformation of Zm401 gene in Lilium longiflorum×L. formosanum." Chinese Journal of Agricultural Biotechnology 5, no. 2 (August 2008): 113–19. http://dx.doi.org/10.1017/s1479236208002222.

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AbstractIn vitro bulb scales of Lilium longiflorum×L. formosanum were used as explants to develop a highly efficient regeneration system. A high regeneration rate (100%) was reached through organogenesis on basal Murashige and Skoog (MS) medium supplemented with 1.0 mg/l 6-benzylaminopurine (6-BA) and 1.0 mg/l naphthaleneacetic acid (NAA). A genetic transformation system for the lily was developed using an Agrobacterium tumefaciens-mediated method. An improved genetic transformation rate (12‰) was obtained when the explants were pre-cultured for 3 days, immersed in bacterial suspension (OD600≈0.8) for 5 min, and co-cultivated for 5 days. The binary vector pBI121 containing Zm401, a maize pollen-specific gene, was introduced into the Agrobacterium strain LBA4404 and transformed into the explants using the genetic transformation system. Gene integration into the lily genome was confirmed by polymerase chain reaction (PCR) and PCR–Southern analysis. These results could lead to the production of new pollenless lily plants.

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Prozorov, A. A. "Milestones of studying the mechanisms of the genetic bacterial transformation on the model of Streptococcus." Biology Bulletin Reviews 6, no. 4 (July 2016): 267–75. http://dx.doi.org/10.1134/s2079086416040046.

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Ma, Xiaolei, Kehou Pan, Lin Zhang, Baohua Zhu, Guanpin Yang, and Xiangyang Zhang. "Genetic transformation of Nannochloropsis oculata with a bacterial phleomycin resistance gene as dominant selective marker." Journal of Ocean University of China 15, no. 2 (February 2, 2016): 351–56. http://dx.doi.org/10.1007/s11802-016-2715-4.

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Nielsen, Kaare M., Kornelia Smalla, and Jan D. van Elsas. "Natural Transformation of Acinetobactersp. Strain BD413 with Cell Lysates of Acinetobacter sp.,Pseudomonas fluorescens, and Burkholderia cepaciain Soil Microcosms." Applied and Environmental Microbiology 66, no. 1 (January 1, 2000): 206–12. http://dx.doi.org/10.1128/aem.66.1.206-212.2000.

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ABSTRACT To elucidate the biological significance of dead bacterial cells in soil to the intra- and interspecies transfer of gene fragments by natural transformation, we have exposed the kanamycin-sensitive recipient Acinetobacter sp. strain BD413(pFG4) to lysates of the kanamycin-resistant donor bacteria Acinetobacterspp., Pseudomonas fluorescens, and Burkholderia cepacia. Detection of gene transfer was facilitated by the recombinational repair of a partially (317 bp) deleted kanamycin resistance gene in the recipient bacterium. The investigation revealed a significant potential of these DNA sources to transformAcinetobacter spp. residing both in sterile and in nonsterile silt loam soil. Heat-treated (80°C, 15 min) cell lysates were capable of transforming strain BD413 after 4 days of incubation in sterile soil and for up to 8 h in nonsterile soil. Transformation efficiencies obtained in vitro and in situ with the various lysates were similar to or exceeded those obtained with conventionally purified DNA. The presence of cell debris did not inhibit transformation in soil, and the debris may protect DNA from rapid biological inactivation. Natural transformation thus providesAcinetobacter spp. with an efficient mechanism to access genetic information from different bacterial species in soil. The relatively short-term biological activity (e.g., transforming activity) of chromosomal DNA in soil contrasts the earlier reported long-term physical stability of DNA, where fractions have been found to persist for several weeks in soil. Thus, there seems to be a clear difference between the physical and the functional significance of chromosomal DNA in soil.

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Ichihara, Hisako, Kei-ichi Kuma, and Hiroyuki Toh. "Positive Selection in the ComC-ComD System of Streptococcal Species." Journal of Bacteriology 188, no. 17 (September 1, 2006): 6429–34. http://dx.doi.org/10.1128/jb.00484-06.

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ABSTRACT Competence-stimulating peptide (CSP) and ComD of the streptococcal species are a pheromone and its receptor, respectively, involved in the regulation of competence for natural genetic transformation. We show here that these molecules have undergone positive selection. This study is the first report of positive selection due to competition among bacterial populations.

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Li, Yaning, Taesik Uhm, Chengwei Ren, Chengcang Wu, Teofila S. Santos, Mi-Kyung Lee, Bo Yan, et al. "A plant-transformation-competent BIBAC/BAC-based map of rice for functional analysis and genetic engineering of its genomic sequence." Genome 50, no. 3 (February 2007): 278–88. http://dx.doi.org/10.1139/g07-006.

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Sequencing of the rice genome has provided a platform for functional genomics research of rice and other cereal species. However, multiple approaches are needed to determine the functions of its genes and sequences and to use the genome sequencing results for genetic improvement of cereal crops. Here, we report a plant-transformation-competent, binary bacterial artificial chromosome (BIBAC) and bacterial artificial chromosome (BAC) based map of rice to facilitate these studies. The map was constructed from 20 835 BIBAC and BAC clones, and consisted of 579 overlapping BIBAC/BAC contigs. To facilitate functional analysis of chromosome 8 genomic sequence and cloning of the genes and QTLs mapped to the chromosome, we anchored the chromosomal contigs to the existing rice genetic maps. The chromosomal map consists of 11 contigs, 59 genetic markers, and 36 sequence tagged sites, spanning a total of ca. 38 Mb in physical length. Comparative analysis between the genetic and physical maps of chromosome 8 showed that there are 3 “hot” and 2 “cold” spots of genetic recombination along the chromosomal arms in addition to the “cold spot” in the centromeric region, suggesting that the sequence component contents of a chromosome may affect its local genetic recombination frequencies. Because of its plant transformability, the BIBAC/BAC map could provide a platform for functional analysis of the rice genome sequence and effective use of the sequencing results for gene and QTL cloning and molecular breeding.

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Rangslang, Rishan Kupar, Ziqi Liu, Henrik Lütken, and Bruno Trevenzoli Favero. "Agrobacterium spp. genes and ORFs: Mechanisms and applications in plant science." Ciência e Agrotecnologia 42, no. 5 (September 2018): 453–63. http://dx.doi.org/10.1590/1413-70542018425000118.

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ABSTRACT The bacterial origin of crown gall tumours, Agrobacterium tumefaciens was isolated 100 years ago. 70 years later, the findings that random integration of bacterial DNA into the host plant genome provided the potential of using Agrobacterium as a plant genetic engineering tool. Since the 1980s, Agrobacterium-mediated transformation on wide range of plants began developing rapidly. This review focused on the oncogenes of A. rhizogenes which is the causative agent of the hairy root disease and the application of A.rhizogenes in plant science.

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Attaiech, Laetitia, Chantal Granadel, Jean-Pierre Claverys, and Bernard Martin. "RadC, a Misleading Name?" Journal of Bacteriology 190, no. 16 (June 13, 2008): 5729–32. http://dx.doi.org/10.1128/jb.00425-08.

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ABSTRACT The pfam04002 annotation describes RadC as a bacterial DNA repair protein. Although the radC gene is expressed specifically during competence for genetic transformation in Streptococcus pneumoniae, we report that radC mutants exhibit normal uptake and processing of transforming DNA. They also display normal sensitivity to DNA-damaging agents, providing no support for the rad epithet.

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Das, Pronabananda, Aneesa Ansari, Mohammad Nurul Islam, and RH Sarker. "Genetic Transformation of a Local Tomato (Solanum lycopersicum L.) Variety of Bangladesh." Plant Tissue Culture and Biotechnology 25, no. 1 (July 9, 2015): 87–97. http://dx.doi.org/10.3329/ptcb.v25i1.24128.

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An in vitro regeneration and Agrobacterium?mediated genetic transformation protocol was optimized for a local tomato variety, BARI Tomato?8 using cotyledonary leaf and hypocotyls explants. The explants were treated with various growth regulators in MS at different concentrations and combinations. Highest number of multiple shoot induction was observed from both the explants cultured in MS supplemented with 8.88 ?M BAP and 0.57 ?M IAA. Half strength of MS supplemented with 1.14 ?M IAA was found to be the best for root induction from excised shoots. Agrobacterium mediated genetic transformation was carried using pBI121 plasmid harboring ??glucuronidase (GUS) reporter and nptII selectable marker genes. Transient GUS assay confirmed that both the explants pre?cultured for two days showed best transformation efficiency in bacterial suspension having optical density (OD) of 0.8 (at 600 nm) for 15 min and co?cultivation period of 3 days. The shoots regenerated from transformed cotyledonary leaf explants survived at 200 mg/l kanamycin selection. The presence of expected amplicon corresponding to the GUS gene was confirmed by PCR. This protocol paves a way for developing disease resistant tomato variety using target gene/s.Plant Tissue Cult. & Biotech. 25(1): 87-97, 2015 (June)

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Agrawal, Shreni, and Esha Rami. "A Review: Agrobacterium-mediated Gene Transformation to Increase Plant Productivity." Journal of Phytopharmacology 11, no. 2 (April 10, 2022): 111–17. http://dx.doi.org/10.31254/phyto.2022.11211.

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In genetics and molecular biology Gene transformation is a gene alteration technique that involves the introduction and expression of a foreign gene into the host organism. There are many gene transformation methods like particle bombardment electroporation micro-injection, (PEG), for different biotechnological experiments But Plant gene transformation is a widely used procedure for obtaining transgenic plants and plant models to understand gene functions. Agrobacterium tumefaciens is a natural genetic engineer which is rod-shaped gram-negative soil-born barteri. Initially Agrobacterium was utilized to transform only dicot plants but over the year’s modification in plant transformation protocol. It was now utilized in monocot plants as well as in fruits plants too. Agrobacterium tumefaciens inserts its (DNA), (Transfer DNA-T-DNA), into the host plant. The transmitted (DNA), is randomly integrated into the host cell's genetic material inside the infected plant cell nucleus. Alternatively bacterial DNA, can transiently remain in the nucleus without integrating into the genome but it still replicates alongside the plant genome, using its machinery and expressing its genes to make separate gene products. Besides the traditional method new research has also been done to transform the plants through agrobacterium. Various methods have been developed to transform monocotyledonous plants such as wheat maize rice and fruity plants. Generally dicotyledonous plants can be transformed by the traditional method of agrobacterium but various methods have also been developed for dicots for various applications. Here, we have taken an example of a tobacco plant (Nicotiana tabacum), transformed with different methods.

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Gulig, Paul A., Matthew S. Tucker, Patrick C. Thiaville, Jennifer L. Joseph, and Roslyn N. Brown. "USER Friendly Cloning Coupled with Chitin-Based Natural Transformation Enables Rapid Mutagenesis of Vibrio vulnificus." Applied and Environmental Microbiology 75, no. 15 (June 5, 2009): 4936–49. http://dx.doi.org/10.1128/aem.02564-08.

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ABSTRACT Vibrio vulnificus is a bacterial contaminant of shellfish and causes highly lethal sepsis and destructive wound infections. A definitive identification of virulence factors using the molecular version of Koch's postulates has been hindered because of difficulties in performing molecular genetic analysis of this opportunistic pathogen. For example, conjugation is required to introduce plasmid DNA, and allelic exchange suicide vectors that rely on sucrose sensitivity for counterselection are not efficient. We therefore incorporated USER friendly cloning techniques into pCVD442-based allelic exchange suicide vectors and other expression vectors to enable the rapid and efficient capture of PCR amplicons. Upstream and downstream DNA sequences flanking genes targeted for deletion were cloned together in a single step. Based on results from Vibrio cholerae, we determined that V. vulnificus becomes naturally transformable with linear DNA during growth on chitin in the form of crab shells. By combining USER friendly cloning and chitin-based transformation, we rapidly and efficiently produced targeted deletions in V. vulnificus, bypassing the need for two-step, suicide vector-mediated allelic exchange. These methods were used to examine the roles of two flagellin loci (flaCDE and flaFBA), the motAB genes, and the cheY-3 gene in motility and to create deletions of rtxC, rtxA1, and fadR. Additionally, chitin-based transformation was useful in moving antibiotic resistance-labeled mutations between V. vulnificus strains by simply coculturing the strains on crab shells. The methods and genetic tools that we developed should be of general use to those performing molecular genetic analysis and manipulation of other gram-negative bacteria.

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Journal articles on the topic 'Bacterial genetic transformation'

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