Biological Sciences
Agrobacterium
Agrobacterium is a genus of bacteria known for its ability to transfer genetic material into plant cells, leading to the formation of crown gall tumors. This process is used in biotechnology to introduce desired genes into plants for agricultural purposes. Agrobacterium-mediated genetic transformation has been a valuable tool in creating genetically modified crops with improved traits such as pest resistance and increased yield.
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12 Key excerpts on "Agrobacterium"
eBook - PDF- Gupta, Rajan Kumar(Authors)
- 2021(Publication Date)
- Daya Publishing House(Publisher)
Chapter 19 Agrobacterium : A Natural Genetic Engineer Ashutosh Bahuguna, Madhuri K. Lily and Koushalya Dangwal * Department of Biotechnology, Modern Institute of Technology, Dhalwala, Rishikesh – 249 201, Uttarakhand ABSTRACT Agrobacterium is a genus of Gram-negative bacteria that uses horizontal gene transfer to cause tumors in plants. Agrobacterium tumefaciens is the most commonly studied species in this genus. Agrobacterium is well known for its ability to transfer DNA from its genome to plant genome, and for this reason it has become an important tool for plant improvement by genetic engineering. Overall species of Agrobacterium can transfer T-DNA to a broad group of plants (both angiosperm and gymnosperm) and other organism. Yet, individual Agrobacterium have a limited host range, the molecular basis for the limited host range is unknown. Keywords : T-DNA, Ti and Ri Plasmid, vir genes, Crown gall, Binary vector. Introduction The genus Agrobacterium is a group of Gram negative soil bacteria found associated with plants ( Figure 19.1 ). Many members of this group cause disease on plants. Infections at wound sites by Agrobacterium tumefaciens cause crown gall tumors on a wide range of plants including most dicots, some monocots, and some gymnosperms. Infections by A. rhizogenes cause hairy root disease. A. vitis causes tumors and necrotic lesions on grape vines This ebook is exclusively for this university only. Cannot be resold/distributed. and is commonly found in the xylem sap of infected plants. Despite the general perception that most of the agrobacteria cause disease, A. radiobacter , is a non-virulent member of this group most often isolated from soil. (Matthysse, 2005) Figure 19.1: Agrobacterium Cells Observed in the Light Microscope Habitat Agrobacteria are usually found in soil in association with roots, tubers, or underground stems. The bacteria also cause tumors from which they can be isolated.
eBook - PDF- María Alvarez(Author)
- 2011(Publication Date)
- IntechOpen(Publisher)
Part 1 Agrobacterium: New Insights into a Natural Engineer 1 Agrobacterium -Mediated Genetic Transformation: History and Progress Minliang Guo * , Xiaowei Bian, Xiao Wu and Meixia Wu College of Bioscience and Biotechnology, Yangzhou University, Jiangsu, P. R. China 1. Introduction Agrobacterium tumefaciens is a Gram-negative soil phytopathogenic bacterium that causes the crown gall disease of dicotyledonous plants, which is characterized by a tumorous phenotype. It induces the tumor by transferring a segment of its Ti plasmid DNA (transferred DNA, or T-DNA) into the host genome and genetically transforming the host. One century has past after A. tumefaciens was firstly identified as the causal agent of crown gall disease (Smith & Townsend, 1907). However, A. tumefaciens is still central to diverse fields of biological and biotechnological research, ranging from its use in plant genetic engineering to representing a model system for studies of a wide variety of biological processes, including bacterial detection of host signaling chemicals, intercellular transfer of macromolecules, importing of nucleoprotein into plant nuclei, and interbacterial chemical signaling via autoinducer-type quorum sensing (McCullen & Binns, 2006; Newton & Fray, 2004; Pitzschke & Hirt, 2010). Therefore, the molecular mechanism underlying the genetic transformation has been the focus of research for a wide spectrum of biologists, from bacteriologists to molecular biologists to botanists. 1.1 History of Agrobacterium tumefaciens research A. tumefaciens is capable of inducing tumors at wound sites of hundreds of dicotyledonous plants, and some monocots and gymnosperms (De Cleene and De Ley, 1976), which may happen on the stems, crowns and roots of the host. At the beginning of the last century, crown gall disease was considered a major problem in horticultural production.
eBook - PDF- Zeb Saddiqe(Author)
- 2019(Publication Date)
- Delve Publishing(Publisher)
The transformation using Agrobacterium has many advantages than methods involving direct gene transfer. For example, it is possible to transfer only a single or limited copies of DNA segments having the genes of choice at high rate with low cost or the transfer of large DNA segments with almost no rearrangements (Shibata and Liu, 2000). The ultimate goal of all these practices is the high yield of a targeted compound, and at the same time reducing the cost as compared to the natural synthesis of that metabolite by the plants. Agrobacterium is a group of widespread naturally occurring soil borne bacteria that infect plants. Infections are caused by the transfer of bacterial genome into the host cells at the point of infection. This bacterial genome then integrates into the genome of host cell. The integrated genetic material is called transferred DNA (T-DNA) that is a part of a large plasmid. A plasmid is a small circular DNA present universally in all bacteria (ABNE, 2010). The inherent ability of these microbes to modify the genetic composition of any plant laid the basis for genetic modification of plants by using Agrobacterium . At present, genetic transformation using Agrobacterium is Medicinal Plants Biotechnology 142 the most widely used procedure for genetic engineering in plants due to its high efficiency. Initially, it was thought that the bacterium only can infect dicot plants, but later it was observed that it can also transform monocot plants for example rice (ABNE, 2010). The genus Agrobacterium has been divided into different species on the basis of disease symptoms and host range. Among different species, A. radiobcater is an “avirulent” species which does not cause any infection. A. rhizogenes causes hairy root disease. A. tumefaciens is the causal organism for crown gall disease in plants. The A. rubi causes cane gall disease while A. vitis infects grapevines (Rout, 2001).
- Prasad, Birendra(Authors)
- 2018(Publication Date)
- Biotech(Publisher)
Chapter 6 Gene Delivery through Agrobacterium Mediated Genetic Transformation in Fruit Crops Mahamadtoufeeq Husainnaik and K.S. Nagaraja KRC College of Horticulture, Arabhavi, Tq. Gokak, Dist. Belgaum -591218, Karnataka Agrobacterium is a soil borne, gram negative, rod shaped, motile found in rhizosphere. It is a causative agents of Crown gall disease of dicotyledones. It is having an ability of transfer of bacterial genes to the plant genome. Agrobacterium will be attracted to wound site via chemotaxis in response to chemicals (sugar and Phenolic molecules: acetosyringone) released from damaged plant cells. This Agrobacterium will contains Ti plasmid which can transfer its T-DNA region into genome of host plants. Agrobacterium genetically transforms its host by transferring a well-defined DNA segment from its tumor-inducing(Ti) plasmid to the host-cell genome. In nature, the transferred DNA (T-DNA) carries a set of oncogenes and opine-catabolism genes, the expression of which, in plant cells, leads to neoplastic growth of the transformed tissue and the production of opines, amino acid derivatives used almost exclusively by the bacteria as a nitrogen source. Recombinant Agrobacterium strains, in which the native T-DNA has been replaced with genes of interests, are the most efficient vehicles used This ebook is exclusively for this university only. Cannot be resold/distributed. today for the introduction of foreign genes into plants and for theproduction of transgenic plant species. Thus, Agrobacterium biology and biotechnology have been the subject of numerous studies over the past few decades, resulting in the establishment of many Agrobacterium strains, plasmids and protocols uniquely adapted for the genetic transformation of various plant species. The molecular machinery needed for T-DNA production and transport into the host cell comprises proteins that are encoded by a set of bacterial chromosomal (chv) and Ti-plasmid virulence (vir) genes.
- Sinha, Asha(Authors)
- 2018(Publication Date)
- Biotech(Publisher)
Chapter 11 Agrobacterium tumefaciens: A Natural Genetic Engineer for Gene Transfer in Plants Jyoti Rastogi 1 , N.N. Tiwari 1 , R.N. Pandey 1 , P. Bubber 2 and R.K. Singh 1 * 1 Center for Sugarcane Biotechnology, U.P. Council of Sugarcane Research, Shahjahanpur — 242 001, U.P. 2 Biochemistry Discipline, School of Sciences, IGNOU, New Delhi —110 068 Globally, Agrobacterium tumefaciens is used as a vector for the development of transgenic plants since few decades. Agrobacterium mediated genetic transformation is the prevailing technology used for the production of genetically modified crop. Some developed countries are using transgenic varieties of economically important crops such as cotton, soybeans, corn, potatoes and tomatoes. This is most efficient method over direct gene transfer approach due to various advantages viz., high transformation efficiency, insertion of single copy number of desired gene, less chances of gene silencing, the insertion of relatively large DNA segment, simple, safe and comparatively less expensive. Agrobacterium tumefaciens is negatively gram stained, soil resident and a member of the alpha-proteobacteria, it can transfer and integrate its oncogenic T-DNA (transfer DNA) from its tumour-inducing (Ti) plasmid into the chromosome of a wide variety of plants (Citovsky et al., 2007). It is bacilliform rod shaped (1x3 pm), motile and belongs to family Rhizobiaceae. This ebook is exclusively for this university only. Cannot be resold/distributed. This bacterium can live in the living state in many soils with good aeration such as sandy loams where crown gall diseased plants have grown. The genus Agrobacterium is divided into a number of species as A. radiobacter is an avirulent species, A. tumefaciens causes crown gall disease, A. rhizogenes causes hairy root disease, and A. rubi causes cane gall disease (Kersters and De Ley, 1984). More recently, a new species has introduced A. vitis which causes galls on grape and a few other plant species.
eBook - PDFGenetic Engineering
An Insight into the Strategies and Applications
- Farrukh Jamal(Author)
- 2016(Publication Date)
- IntechOpen(Publisher)
In general, the rate of transformed cells in tissue is quite low. That is why the prerequisite of success in gene transfer is high-frequency shoot regeneration. Most commonly used technique in gene transfer to plants is the bacterium Agrobacterium tumefaciens . A . tumefaciens is known as a “natural genetic engineer of plants” due to this trait [ 1]. Agrobacterium -mediated transforma-tion method has been a widely used gene transfer method. The advantages of the method are wide host range of plants: agronomically and horticulturally important crops including soy-bean, cotton, rice, wheat, flowers, and various trees [ 2] and transferring a small copy number of the transfer-DNA (T-DNA) into the cytoplasm and resulting in stable integration into plant chromosome. Although it has merits as compared to other transformation methods, such as particle bombardment, electroporation, and silicon carbide fibers, it is still hard to achieve high transformation efficiency and gene expression using this method. 2. Molecular mechanism of A . tumefaciens -mediated DNA transfer Agrobacterium , of the family Rhizobiaceae , is a genus of Gram-negative bacterium that geneti-cally transforms host plants and causes crown gall tumors at wound sites [ 3] ( Figure 1 ). Agrobacterium can transfer DNA to a broad group of organisms: plants, fungi such as yeasts, ascomycetes, and basidiomycetes, and protist such as algae [ 4, 5]. Agrobacterium is usually clas-sified by the disease symptomology (type of opine) and host range. The genetic mechanism of Figure 1. Crown gall in sugar beet caused by wild (oncogenic) Agrobacterium strain. Genetic Engineering - An Insight into the Strategies and Applications 24 host range determination is still obscure, but it was reported that several virulence ( vir ) genes on the tumor-inducing (Ti) plasmid, virC [ 6], virF [ 7], and virH [ 8] were involved in determi-nation for the range of plant species.
eBook - ePub- Alan R. Lindsey(Author)
- 2022(Publication Date)
- Routledge(Publisher)
This chapter starts with an overview of the current knowledge on the molecular basis of Agrobacterium -mediated plant transformation. The main emphasis is on the role of the virulence proteins and on recent insights into the later steps of the transformation process. Then, a summary is given on the characteristics of the T-DNA inserted into the plant nuclear DNA. The chapter concludes with a critical assessment of the different approaches that have been used to exploit Agrobacterium tumefaciens, a natural “engineer”, for the production of transgenic plants. Key words: Agrobacterium tumefaciens, plant transformation, regeneration, selectable marker, T-DNA, virulence. Introduction The dawn of plant molecular biology and genetic engineering was basically the consequence of the discovery and study of the plant pathogen Agrobacterium tumefaciens. In nature, this Gram-negative soil bacterium causes tumour formation (so-called crown galls) on a large number of dicotyledonous as well as some monocotyledonous plant species and Gymnosperms (De Cleene and De Ley, 1976). These tumours can be grown in vitro in simple media without the continued presence of the bacterium (Braun and White, 1943). Although A. tumefaciens had been studied for nearly 100 years, it was the advent of recombinant DNA technology that led to the analysis of the biological principles underlying the Agrobacterium –plant cell interaction and to the modification of the system for plant genetic engineering. The crown gall disease has been shown to be due to the transfer of a specific DNA fragment, the T-DNA (transferred DNA), from a large tumour-inducing (Ti) plasmid within the bacterium (Zaenen et al., 1974) to the plant cell (summarized in Figure 1.1). After transfer, the T-DNA becomes integrated into the plant nuclear genome (Chilton et al., 1977; Schell et al., 1979) and its subsequent expression leads to the crown gall phenotype
eBook - ePub- Jayarama Reddy(Author)
- 2024(Publication Date)
- CRC Press(Publisher)
virG, contribute to the “hypervirulence” of particular strains.A. tumefaciens Causal Agent of Crown Gall DiseaseCrown Gall disease caused by AgrobacteriumMolecular Basis of Agrobacterium–Mediated Transformation T-DNA
The molecular basis of genetic transformation of plant cells by Agrobacterium is transfer from the bacterium and integration into the plant nuclear genome of a region of a large tumor-inducing (Ti) or rhizogenic (Ri) plasmid resident in Agrobacterium. Ti plasmids are on the order of 200 to 800 kbp in size. The transferred DNA (T-DNA) or Ri plasmid. T-regions on native Ti and Ri plasmids are approximately 10 to 30 kbp in size. Thus, T-regions generally represent less than 10% of the Ti plasmid. Some Ti plasmids contain one T-region, whereas others contain multiple T-regions. The processing of the T-DNA from the Ti plasmid and its subsequent export from the bacterium to the plant cell result in large part from the activity of virulence (vir) genes carried by the Ti plasmid. T-regions are defined by T-DNA border sequences. These borders are 25 bp in length and highly homologous in sequence. They flank the T-region in a directly repeated orientation. In general, the T-DNA borders delimit the T-DNA, because these sequences are the target of the VirD1/VirD2 borderspecific endonuclease that processes the T-DNA from the Ti plasmid. There appears to be a polarity established among T-DNA borders: right borders initially appeared to be more important than left borders. We now know that this polarity may be caused by several factors. First, the border sequences not only serve as the target for the VirD1/VirD2 endonuclease but also serve as the covalent attachment site for VirD2 protein. Within the Ti or Ri plasmid (or T-DNA binary vectors), T-DNA borders are made up of double- stranded DNA. Cleavage of these double stranded border sequences requires VirD1 and VirD2 proteins, both in vivo and in vitro. In vitro, however, VirD2 protein alone can cleave a single- stranded T-DNA border sequence. Cleavage of the 25-bp T-DNA border results predominantly from the nicking of the T-DNA “lower strand,” as conventionally presented, between nucleotides 3 and 4 of the border sequence. However, double-strand cleavage of the T-DNA border has also been noted. Nicking of the border is associated with the tight (probably covalent) linkage of the VirD2 protein, through tyrosine 29, to the 5’ end of the resulting single stranded T-DNA molecule termed the T-strand. It is ssT-strand, and not a double- stranded T-DNA molecule, that is transferred to the plant cell. Thus, it is the VirD2 protein attached to the right border and not the border sequence per se, that establishes polarity and the importance of right borders relative to left borders. It should be noted, however, that because left-border nicking is also associated with VirD2 attachment to the remaining molecule (the “non-T- DNA” portion of the Ti plasmid or “backbone” region of the T- DNA binary vector), it may be possible to process T-strands from these regions of Ti and Ri plasmids and from T-DNA binary vectors. Second, the presence of T-DNA “overdrive” sequences near many T-DNA right borders, but not left borders, may also help establish the functional polarity of right and left borders. Overdrive sequences enhance the transmission of T-strands to plants, although the molecular mechanism of how this occurs remains unknown. Early reports suggested that the VirC1 protein binds to the overdrive sequence and may enhance T-DNA border cleavage by the VirD1/VirD2 endonuclease. VirC1 and virC2
eBook - PDF- Behera, K.K.(Authors)
- 2020(Publication Date)
- NEW INDIA PUBLISHING AGENCY (NIPA)(Publisher)
Contact Between Plant Cells and Bacterial Cells Wounding of a plant, which often occurs in the region of the stem-root interface, allow the entry of bacterium. Bacteria are attracted to the wounded plant in response to signal molecules that are released by the plant cells; they multiply in the wound sap and attach to the walls of plant cells. The specific attachment of Agrobacterium to plant cells is a prerequisite for subsequent transfer of DNA. Products of several chromosomal genes ( chv A, chv B, pac A and att ) in Agrobacterium and surface receptors of plant cells, which might include both protein and carbohydrates, are believed to be involved in this process. Activation of Virulence ( vir ) Genes The T-DNA fragment is flanked by 25-bp direct repeats, which act in cis as a signal for the transfer apparatus. The process of T-DNA transfer is mediated by the co-operative action of proteins encoded by genes of the Ti plasmid virulence region (vir genes) and in the bacterial genome. Following transfer and expression of the T-DNA into the plant nucleus, the plant cells proliferate and synthesize certain types of opines, depending upon the T-DNA type that has been integrated. These opines in turn can serve as a sole C and N source for Agrobacterium . The 30 kb vir region is a regulon organised of seven operons that are essential for T-DNA transfer ( virA, virB, virD, and virG ) and for the increasing of transfer efficiency ( virC and virE ). Several genes of the bacterial chromosome have been shown to function in the attachment of Agrobacterium , to the plant cell and in bacterial colonisation.
eBook - PDF- David A. Hopwood, Keith F. Chater(Authors)
- 2013(Publication Date)
- Academic Press(Publisher)
Section Bacteria that Interact with Plants as Parasites or Symbionts Members of a group of related genera in the a subdivision of the purple bacteria have evolved special relationships with higher plants in which plant tissue is induced to proliferate to give rise to nodules or benign tumours which become the habitat for specialized derivatives of the bacteria or some of their genes. One of the recent successes of bacterial genetics has been the under-standing of the roles of genes carried by Agrobacterium species or rhizobia (including members of the new genera Rhizobium, Bradyrhizobium and Azorhizobium) in the genetic transformation of plant tissue in crown gall or hairy root disease on the one hand, or in root nodule formation and nitrogen fixation on the other (Chapters 18 and 19). While bacterial chromosomal genes undoubtedly play a role in determining the outcome of the interaction between the bacterium and its host plant (those for exopolysaccharide produc-tion by rhizobia being examples), remarkably, the genes that determine the formation of galls or root nodules, as well as the specialized biochemical functions manifested in them —opine and plant hormone production, or nitrogen fixation, respectively —all form parts of large plasmids whose posses-sion by a relatively unspecialized Gram-negative rod converts it into a unique plant parasite or symbiont. The molecular understanding of crown gall was greatly simplified by the circumscribed nature of the disease. Other bacterial diseases of plants approach in complexity those of animals described in Section IV and are correspondingly difficult to understand. In vertebrates, specific responses of the immune system to infection may often help to pinpoint particular attributes of the pathogen that are likely to be relevant to the development of disease. The V 351
eBook - PDF- David A. Hopwood, Keith F. Chater(Authors)
- 2012(Publication Date)
- Academic Press(Publisher)
Section Bacteria that Interact with Plants as Parasites or Symbionts Members of a group of related genera in the a subdivision of the purple bacteria have evolved special relationships with higher plants in which plant tissue is induced to proliferate to give rise to nodules or benign tumours which become the habitat for specialized derivatives of the bacteria or some of their genes. One of the recent successes of bacterial genetics has been the under-standing of the roles of genes carried by Agrobacterium species or rhizobia (including members of the new genera Rhizobium, Bradyrhizobium and Azorhizobium) in the genetic transformation of plant tissue in crown gall or hairy root disease on the one hand, or in root nodule formation and nitrogen fixation on the other (Chapters 18 and 19). While bacterial chromosomal genes undoubtedly play a role in determining the outcome of the interaction between the bacterium and its host plant (those for exopolysaccharide produc-tion by rhizobia being examples), remarkably, the genes that determine the formation of galls or root nodules, as well as the specialized biochemical functions manifested in them—opine and plant hormone production, or nitrogen fixation, respectively—all form parts of large plasmids whose posses-sion by a relatively unspecialized Gram-negative rod converts it into a unique plant parasite or symbiont. The molecular understanding of crown gall was greatly simplified by the circumscribed nature of the disease. Other bacterial diseases of plants approach in complexity those of animals described in Section IV and are correspondingly difficult to understand. In vertebrates, specific responses of the immune system to infection may often help to pinpoint particular attributes of the pathogen that are likely to be relevant to the development of disease. The v 351
eBook - PDFTransgenic Plants
Advances and Limitations
- Yelda and #214;zden and #199;ift and #231;i(Author)
- 2012(Publication Date)
- IntechOpen(Publisher)
Part 1 Application 1 Agrobacterium -Mediated Transformation of Wheat: General Overview and New Approaches to Model and Identify the Key Factors Involved Pelayo Pérez-Piñeiro 1 , Jorge Gago 1 , Mariana Landín 2 and Pedro P. . Gallego 1,* 1 Applied Plant and Soil Biology, Dpt. Plant Biology and Soil Science, Faculty of Biology, University of Vigo,Vigo, 2 Dpt. Pharmacy and Pharmaceutical Technology, Faculty of Pharmacy, University of Santiago, Santiago de Compostela, Spain 1. Introduction Wheat is the world’s second largest crop, supplying 19% of human calories; the largest volume crop traded internationally and grown on approximately 17% of the world’s cultivatable land (over 200 million hectares) (Jones, 2005; Atchison et al., 2010). However, probably due to climate change, some adverse environmental conditions have caused a downward trend in world wheat production (FAO, 2003; 2011). In this context, developing new higher yielding wheat varieties more tolerant or resistant to abiotic and/or biotic stress, using all available plant biotechnology technologies available, should be considered as the major challenge. The scientific community has made considerable efforts to understand and improve the goal of the integration of an exogenous T-DNA in the genome of a host plant cell and, subsequently, the regeneration into a whole plant. The most extended method for plant genetic transformation uses the Agrobacterium bacteria as the biological vector to transfer exogenous T-DNA into the plant cell. Although, Agrobacterium -mediated transformation became widely available for the routine transformation of most crops, cereals initially have been recalcitrant to this system, since these crops were not naturally susceptible to Agrobacterium sp (Potrykus, 1990, 1991).
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