View source for Biology:Ti plasmid

[[File:Ti plasmid.svg|thumb|350px|right|alt=A diagram showing the structure of the Ti plasmid, with various important regions labeled|The structure of the Ti plasmid]]
A '''tumour inducing (Ti)''' [[Biology:Plasmid|plasmid]] is a plasmid found in pathogenic species of ''[[Biology:Agrobacterium|Agrobacterium]]'', including [[Biology:Agrobacterium tumefaciens|''A. tumefaciens]], [[Biology:Rhizobium rhizogenes|''A. rhizogenes'']], ''A. rubi'' and [[Biology:Allorhizobium vitis|''A. vitis'']].

Evolutionarily, the Ti plasmid is part of a family of plasmids carried by many species of [[Biology:Alphaproteobacteria|Alphaproteobacteria]]. Members of this plasmid family are defined by the presence of a conserved DNA region known as the ''repABC'' gene cassette, which mediates the [[Biology:DNA replication|replication]] of the plasmid, the partitioning of the plasmid into daughter cells during [[Biology:Cell division|cell division]] as well as the maintenance of the plasmid at low copy numbers in a cell.<ref name="Gordon2014">{{cite journal | vauthors = Gordon JE, Christie PJ | title = The Agrobacterium Ti Plasmids | journal = Microbiology Spectrum | volume = 2 | issue = 6 | date = December 2014 | pmid = 25593788 | pmc = 4292801 | doi = 10.1128/microbiolspec.PLAS-0010-2013 | doi-access = free }}</ref> The Ti plasmids themselves are sorted into different categories based on the type of molecule, or [[Chemistry:Opine|opine]], they allow the bacteria to break down as an energy source.<ref name="Hooykaas1994">{{cite journal | vauthors = Hooykaas PJ, Beijersbergen AG |title=The virulence system of ''Agrobacterium tumefaciens'' |journal=Annual Review of Phytopathology |date=1994 |volume=32 |issue=1 |pages=157–181 |doi=10.1146/annurev.py.32.090194.001105}}</ref>

The presence of this Ti plasmid is essential for the bacteria to cause crown gall disease in plants.<ref name="Gordon2014" /> This is facilitated via certain crucial regions in the Ti plasmid, including the ''vir'' region, which encodes for virulence genes, and the [[Biology:Transfer DNA|transfer DNA]] (T-DNA) region, which is a section of the Ti plasmid that is [[Biology:Horizontal gene transfer|transferred]] via [[Biology:Bacterial conjugation|conjugation]] into host plant cells after an injury site is sensed by the bacteria. These regions have features that allow the delivery of T-DNA into host plant cells, and can modify the host plant cell to cause the synthesis of molecules like plant hormones (e.g. auxins, cytokinins) and opines and the formation of crown gall tumours.<ref name="Gordon2014" />

Because the T-DNA region of the Ti plasmid can be transferred from bacteria to plant cells, it represented an exciting avenue for the transfer of DNA between [[Biology:Kingdom|kingdoms]] and spurred large amounts of research on the Ti plasmid and its possible uses in bioengineering.

==Nomenclature and classification==
The Ti plasmid is a member of the RepABC plasmid family found in Alphaproteobacteria.<ref name="Pinto2012">{{cite journal | vauthors = Pinto UM, Pappas KM, Winans SC | title = The ABCs of plasmid replication and segregation | journal = Nature Reviews. Microbiology | volume = 10 | issue = 11 | pages = 755–65 | date = November 2012 | pmid = 23070556 | doi = 10.1038/nrmicro2882 | s2cid = 6518175 }}</ref> These plasmids are often relatively large in size, ranging from 100kbp to 2Mbp. They are also often termed [[Biology:Replicon (genetics)|replicons]], as their replication begins at a single site. Members of this family have a characteristic ''repABC'' gene cassette.<ref name="Cevallos2008">{{cite journal | vauthors = Cevallos MA, Cervantes-Rivera R, Gutiérrez-Ríos RM | title = The repABC plasmid family | journal = Plasmid | volume = 60 | issue = 1 | pages = 19–37 | date = July 2008 | pmid = 18433868 | doi = 10.1016/j.plasmid.2008.03.001 }}</ref> Another notable member of this family is the root inducing (Ri) plasmid carried by ''A. rhizogenes'', which causes another plant disease known as hairy root disease.<ref name="Gordon2014" />

A key feature of Ti plasmids is their ability to drive the production of opines, which are derivatives of various amino acids or [[Biology:Sugar phosphates|sugar phosphates]], in host plant cells. These opines can then be used as a nutrient for the infecting bacteria, which [[Biology:Catabolism|catabolizes]] the respective opines using genes encoded in the Ti plasmid.

Accordingly, Ti plasmids have been classified based on the type of opine they catabolize, namely: [[Chemistry:Nopaline|nopaline-]], [[Chemistry:Octopine|octopine-]] or mannityl-types, which are amino acid derivatives, or agrocinopine-type, which are sugar phosphate derivatives.<ref name="Gordon2014" />

==Historical discovery==
The identification of ''A. tumefaciens'' as the cause of gall tumours in plants paved the way for insights into the molecular basis of crown gall disease.<ref name="Kado">{{cite journal | vauthors = Kado CI | title = Historical account on gaining insights on the mechanism of crown gall tumorigenesis induced by Agrobacterium tumefaciens | journal = Frontiers in Microbiology | volume = 5 | issue = 340 | pages = 340 | date = 2014 | pmid = 25147542 | pmc = 4124706 | doi = 10.3389/fmicb.2014.00340 | doi-access = free }}</ref>

The first indication of a genetic effect on host plant cells came in 1942-1943, where plant cells of secondary tumours were found to lack any bacterial cells within. However, these tumour cells did possess the ability to produce opines metabolized by the infecting bacterial strain.<ref>{{cite journal | vauthors = Petit A, Delhaye S, Tempé J, Morel G |title=Recherches sur les guanidines des tissues de Crown gall. Mise en evidence d'une relation biochimique spécifique entre les souches d'''Agrobacterium tumefaciens'' et les tumeurs qu'elles induisent |journal=Physiol. Vég. |date=1970 |volume=8 |pages=205–213 }}</ref> Crucially, the production of the respective opines occurred regardless of the plant species and occasionally only within crown gall tissues, indicating that the bacteria had transferred some genetic material to the host plant cells in order to allow opine synthesis.<ref name="Kado" />

However, how and to what extend did DNA transfer occur remained an open question. Adding ''A. tumefaciens'' DNA alone did not cause tumors in plants,<ref>{{cite journal | vauthors = Kado CI, Lurquin PF |title=Studies on ''Agrobacterium tumefaciens''. V. Fate of exogenously added bacterial DNA in ''Nicotiana tabacum'' |journal=Physiological Plant Pathology |date=1976 |volume=8 |issue=1 |pages=73–82 |doi=10.1016/0048-4059(76)90009-6}}</ref> while very little ''A. tumefaciens'' DNA was found to be integrated into the host plant cell genome.<ref>{{cite journal | vauthors = Drlicá KA, Kado CI | title = Quantitative estimation of Agrobacterium tumefaciens DNA in crown gall tumor cells | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 71 | issue = 9 | pages = 3677–81 | date = September 1974 | pmid = 4530329 | pmc = 433839 | doi = 10.1073/pnas.71.9.3677 | doi-access = free | bibcode = 1974PNAS...71.3677D }}</ref> The addition of deoxyribonucleases (DNases) to degrade DNA also failed to prevent the formation and growth of the plant tumors.<ref>{{cite journal | vauthors = Braun AC, Wood HN | title = On the inhibition of tumor inception in the crown-gall disease with the use of ribonuclease A | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 56 | issue = 5 | pages = 1417–22 | date = November 1966 | pmid = 5230302 | pmc = 219988 | doi = 10.1073/pnas.56.5.1417 | doi-access = free | bibcode = 1966PNAS...56.1417B }}</ref> These suggested that little, if any, of the ''A. tumefaciens'' DNA is transferred to the host plant cell to cause disease and, if DNA is indeed transferred from the bacteria to the plant, it must occur in a protected manner.

Subsequently, oncogenic bacterial strains were found to be able to convert non-pathogenic bacteria into pathogens via the process of conjugation, where the genes responsible for virulence were transferred to the non-pathogenic cells.<ref>{{cite journal | vauthors = Kerr A |title=Acquisition of virulence by non-pathogenic isolates of ''Agrobacterium radiobacter''. |journal=Physiological Plant Pathology |date=1971 |volume=1 |issue=3 |pages=241–246 |doi=10.1016/0048-4059(71)90045-2}}</ref> The role of a plasmid in this pathogenic ability was further supported when large plasmids were found only in pathogenic bacteria but not avirulent bacteria.<ref>{{cite journal | vauthors = Zaenen I, Van Larebeke N, Van Montagu M, Schell J | title = Supercoiled circular DNA in crown-gall inducing Agrobacterium strains | journal = Journal of Molecular Biology | volume = 86 | issue = 1 | pages = 109–27 | date = June 1974 | pmid = 4854526 | doi = 10.1016/s0022-2836(74)80011-2 }}</ref> Eventually, the detection of parts of bacterial plasmids in host plant cells was established, confirming that this was the genetic material responsible for the genetic effect of infection.<ref>{{cite journal | vauthors = Chilton MD, Drummond MH, Merio DJ, Sciaky D, Montoya AL, Gordon MP, Nester EW | title = Stable incorporation of plasmid DNA into higher plant cells: the molecular basis of crown gall tumorigenesis | journal = Cell | volume = 11 | issue = 2 | pages = 263–71 | date = June 1977 | pmid = 890735 | doi = 10.1016/0092-8674(77)90043-5 | s2cid = 7533482 }}</ref>

With the identification of the Ti plasmid, many studies were carried out to determine the characteristics of the Ti plasmid and how the genetic material is transferred from the ''Agrobacterium'' to the plant host. Some notable early milestones in the studies of Ti plasmids include the mapping of a Ti plasmid in 1978 and the studying of sequence similarity between different Ti plasmids in 1981.<ref>{{cite journal | vauthors = Chilton MD, Montoya AL, Merlo DJ, Drummond MH, Nutter R, Gordon MP, Nester EW | title = Restriction endonuclease mapping of a plasmid that confers oncogenicity upon Agrobacterium tumefaciens strain B6-806 | journal = Plasmid | volume = 1 | issue = 2 | pages = 254–69 | date = February 1978 | pmid = 748950 | doi = 10.1016/0147-619x(78)90043-4 }}</ref><ref>{{cite journal | vauthors = Engler G, Depicker A, Maenhaut R, Villarroel R, Van Montagu M, Schell J | title = Physical mapping of DNA base sequence homologies between an octopine and a nopaline Ti plasmid of Agrobacterium tumefaciens | journal = Journal of Molecular Biology | volume = 152 | issue = 2 | pages = 183–208 | date = October 1981 | pmid = 6276566 | doi = 10.1016/0022-2836(81)90239-4 }}</ref>

Between 1980–2000, the characterization of the T-DNA region and the 'vir' region was also pursued. Studies into the T-DNA region determined their process of transfer and identified genes allowing the synthesis of plant hormones and opines.<ref>{{cite journal | vauthors = Zambryski P, Tempe J, Schell J | title = Transfer and function of T-DNA genes from agrobacterium Ti and Ri plasmids in plants | journal = Cell | volume = 56 | issue = 2 | pages = 193–201 | date = January 1989 | pmid = 2643473 | doi = 10.1016/0092-8674(89)90892-1 | s2cid = 19393909 }}</ref> Separately, early work aimed to determine the functions of the genes encoded in the 'vir' region - these were broadly categorized into those that allowed bacterial-host interactions and those that enabled T-DNA delivery.<ref name="Hooykaas1994" />

==Replication, partitioning and maintenance==
[[File:RepABC gene cassette of Ti plasmids.png|thumb|center|upright=1.5|alt=A schematic of the ''repABC'' gene cassette, along with the activity of their gene products|The ''repABC gene'' cassette of Ti plasmids in Agrobacteria, with a schematic of their gene product and activities]]

The replication, partitioning and maintenance of the Ti plasmid depends on the ''repABC'' gene cassette, which is mainly made up of three genes: ''repA'', ''repB'' and ''repC''. ''repA'' and ''repB'' each encode for proteins involved in plasmid partitioning, while ''repC'' encodes a replication initiator.<ref name="Gordon2014" /> These genes are expressed from 4 different promoters located upstream of ''repA''. ''repE'' encodes for a small [[Biology:Antisense RNA|antisense RNA]] and is located between ''repB'' and ''repC''.<ref name="Cevallos2008" /> Additionally, there is a partitioning site (''parS'') and an [[Biology:Origin of replication|origin of replication]] (''oriV'') present within the ''repABC'' cassette.<ref name="Gordon2014" />

===Replication of the Ti plasmid===
The replication of the Ti plasmid is driven by the RepC initiator protein ({{UniProt|P05684}}), which possesses two protein domains: an N-terminal domain (NTD) that binds to DNA and a C-terminal domain (CTD). Mutational analyses have shown that without a functional RepC protein, the Ti plasmid is unable to replicate.<ref name="Cevallos2008" /> Meanwhile, the ''oriV'' sequence is around 150 nucleotides in length and is found within the ''repC'' gene.<ref name="Pinto2012" /> Laboratory experiments have shown that the RepC protein binds to this region, suggesting its role as the origin of replication.<ref name="Pinto2011">{{cite journal | vauthors = Pinto UM, Flores-Mireles AL, Costa ED, Winans SC | title = RepC protein of the octopine-type Ti plasmid binds to the probable origin of replication within repC and functions only in cis | journal = Molecular Microbiology | volume = 81 | issue = 6 | pages = 1593–606 | date = September 2011 | pmid = 21883520 | doi = 10.1111/j.1365-2958.2011.07789.x | doi-access = free }}</ref> Therefore, while the complete process behind the replication of the Ti plasmid has not been fully described, the initial step of replication would likely depend on the expression of RepC and its binding to ''oriV''. Of note, the RepC protein only acts in ''cis'', where it only drives the replication of the plasmid it is encoded in and not any other plasmid also present in the bacterial cell.<ref name="Pinto2011" />

===Partitioning of the Ti plasmid===

{| class="wikitable"
|+ style="text-align:left;" | Components involved in the RepA/RepB partitioning system of Ti plasmids<ref name="Gordon2014" />
|- 
! Component
! Function
|-
| RepA (ParA), {{UniProt|P05682}}
| A weak [[Biology:ATPase|ATPase]] that negatively [[Biology:Autoregulation|autoregulates]] the expression of the ''repABC'' cassette and can form filaments to aid in the partitioning of the plasmid during cell division
|-
| RepB (ParB), {{UniProt|P05683}}
| A DNA binding protein that serves as an adaptor between RepA and the ''parS'' site
|- 
| ''parS''
| The palindromic binding site for the ParB protein; consensus {{DNA sequence|GTTNNCNGCNGNNAAC}}
|}

The [[Biology:Plasmid partition system|partitioning system]] of the Ti plasmid is similar to the ParA/ParB system used in other plasmids and bacterial chromosomes and is thought to act in the same way.<ref name="Bignell2001">{{cite journal | vauthors = Bignell C, Thomas CM | title = The bacterial ParA-ParB partitioning proteins | journal = Journal of Biotechnology | volume = 91 | issue = 1 | pages = 1–34 | date = September 2001 | pmid = 11522360 | doi = 10.1016/S0168-1656(01)00293-0 }}</ref> Mutations in either of the proteins RepA or RepB have resulted in a decrease in plasmid stability, indicating their role and importance in plasmid partitioning.<ref name="Cevallos2008" />  The ability of RepA to form filaments allows it to create a physical bridge along which DNA can be pulled to opposite poles of a dividing cell. Meanwhile, the RepB protein can bind specifically to the ''parS'' sequence, forming a complex with DNA that can be recognized by RepA.<ref name="Gordon2014" /><ref name="Cevallos2008" /> This system is particularly important for the proper partitioning of the Ti plasmid, as the plasmid is only present in few copy numbers in the bacterial cell.

===Maintenance of the Ti plasmid===
The Ti plasmid is maintained at low copy numbers within a bacterial cell. This is partly achieved by influencing the expression of the replication initiator RepC.<ref name="Gordon2014" /> When bound to [[Chemistry:Adenosine diphosphate|ADP]], RepA is activated to work with RepB, acting as a negative [[Biology:Regulation of gene expression|regulator]] of the ''repABC'' cassette.<ref name="Pinto2012" /> The levels of RepC is therefore kept low within a cell, preventing too many rounds of replication from occurring during each cell division cycle. Furthermore, there is a small RNA known as RepE encoded between ''repB'' and ''repC'' that lowers the expression of ''repC''.<ref name="Chai2005">{{cite journal | vauthors = Chai Y, Winans SC | title = A small antisense RNA downregulates expression of an essential replicase protein of an Agrobacterium tumefaciens Ti plasmid | journal = Molecular Microbiology | volume = 56 | issue = 6 | pages = 1574–85 | date = June 2005 | pmid = 15916607 | doi = 10.1111/j.1365-2958.2005.04636.x | s2cid = 20348880 | doi-access = free }}</ref> RepE is [[Biology:Complementarity (molecular biology)|complementary]] to RepC and will bind with the ''repC'' [[Biology:Messenger RNA|mRNA]] to form a double-stranded molecule. This can then block the [[Biology:Translation|translational]] production of the RepC protein.<ref name="Chai2005" />

Separately, the expression of the ''repABC'' cassette and hence the copy number of the Ti plasmid is also influenced via a [[Biology:Quorum sensing|quorum sensing]] system in ''Agrobacterium''.<ref name="Cevallos2008" /> Quorum sensing systems respond to bacterial population densities by sensing a molecule, known as an autoinducer, that is produced by the bacterial cells at low levels and would build up to a threshold level when there is a high density of bacteria present.<ref name="Chai2005"/> In this case, the autoinducer is the N-3-oxooctanoyl-L-homoserine lactone (3-O-C<sub>8</sub>-AHL) molecule, which is sensed by a regulator known as TraR.<ref name="Cevallos2008" /> When activated, TraR will bind to regions known as ''tra'' boxes in the ''repABC'' gene cassette's promoter regions to drive expression. Therefore, a high level of population density increases the number of plasmids present within each bacterial cell, likely to support pathogenesis in the plant host.<ref name="Cevallos2008" />

==Features==
{{missing information|section|UniProt links for easier exploration|date=December 2020}}

===Virulence operon===
[[File:Vir region of Ti plasmids.png|thumb|center|upright=2.0|alt=A diagram showing the composition of the ''vir'' region of Ti plasmids|The composition of the ''vir'' region of octopine-type Ti plasmids]]

The expression of the ''vir'' region is usually repressed under normal conditions, and only becomes activated when the bacteria senses plant-derived signals from wound sites. This activation is necessary for the production of Vir proteins and the transfer of DNA and proteins into host plant cells.<ref name="Gordon2014" />

VirA and VirG form a [[Biology:Two-component regulatory system|two-component regulatory system]] within ''Agrobacterium''.<ref name="Winans1991">{{cite journal | vauthors = Winans SC | title = An Agrobacterium two-component regulatory system for the detection of chemicals released from plant wounds | journal = Molecular Microbiology | volume = 5 | issue = 10 | pages = 2345–50 | date = October 1991 | pmid = 1791750 | doi = 10.1111/j.1365-2958.1991.tb02080.x | s2cid = 21209713 }}</ref> This is a type of sensing and signalling system found commonly in bacteria; in this case, they act to sense plant-derived signals to drive the expression of the ''vir'' region. During the sensing, VirA, a histidine sensor kinase, will become [[Biology:Phosphorylation|phosphorylated]] before passing on this phosphate group to the response regulator VirG.<ref>{{cite journal | vauthors = Huang Y, Morel P, Powell B, Kado CI | title = VirA, a coregulator of Ti-specified virulence genes, is phosphorylated in vitro | journal = Journal of Bacteriology | volume = 172 | issue = 2 | pages = 1142–4 | date = February 1990 | pmid = 2298696 | pmc = 208549 | doi = 10.1128/jb.172.2.1142-1144.1990 | doi-access = free }}</ref> The activated response regulator VirG can then bind to a region of DNA known as the ''vir'' box, located upstream of each ''vir'' promoter, to activate the expression of the ''vir'' region.<ref name="Gordon2014" /><ref name="Winans1991" /> One possible downstream functions of the sensing mediated by VirA and VirG is the directional movement, or [[Physics:Chemotaxis|chemotaxis]], of the bacteria towards plant-derived signals; this allows the ''Agrobacterium'' to move towards the wound site in plants.<ref>{{cite journal | vauthors = Shaw CH, Ashby AM, Brown A, Royal C, Loake GJ, Shaw CH | title = virA and virG are the Ti-plasmid functions required for chemotaxis of Agrobacterium tumefaciens towards acetosyringone | journal = Molecular Microbiology | volume = 2 | issue = 3 | pages = 413–7 | date = May 1988 | pmid = 3398775 | doi = 10.1111/j.1365-2958.1988.tb00046.x | s2cid = 8536451 }}</ref> Furthermore, with the induction of the ''vir'' region, the transfer of T-DNA can be mediated by the Vir proteins.<ref>{{cite journal | vauthors = Stachel SE, Zambryski PC | title = virA and virG control the plant-induced activation of the T-DNA transfer process of A. tumefaciens | journal = Cell | volume = 46 | issue = 3 | pages = 325–33 | date = August 1986 | pmid = 3731272 | doi = 10.1016/0092-8674(86)90653-7 | s2cid = 37938846 }}</ref>

The ''virB'' [[Biology:Operon|operon]] is the largest operon in the ''vir'' region, encoding for 11 VirB proteins involved in the transfer process of T-DNA and bacterial proteins into host plant cells (see transfer apparatus below).<ref>{{cite journal | vauthors = Ward JE, Akiyoshi DE, Regier D, Datta A, Gordon MP, Nester EW | title = Characterization of the virB operon from an Agrobacterium tumefaciens Ti plasmid | journal = The Journal of Biological Chemistry | volume = 263 | issue = 12 | pages = 5804–14 | date = April 1988 | doi = 10.1016/S0021-9258(18)60637-4 | pmid = 3281947 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Vergunst AC, Schrammeijer B, den Dulk-Ras A, de Vlaam CM, Regensburg-Tuïnk TJ, Hooykaas PJ | title = VirB/D4-dependent protein translocation from Agrobacterium into plant cells | journal = Science | volume = 290 | issue = 5493 | pages = 979–82 | date = November 2000 | pmid = 11062129 | doi = 10.1126/science.290.5493.979 | bibcode = 2000Sci...290..979V }}</ref>

The ''virC'' operon encodes for two proteins: VirC1 and VirC2. These proteins influence the pathogenesis of the ''Agrobacterium'' towards different plant hosts, and mutations can reduce but not remove the virulence of the bacteria.<ref>{{cite journal | vauthors = Close TJ, Tait RC, Rempel HC, Hirooka T, Kim L, Kado CI | title = Molecular characterization of the virC genes of the Ti plasmid | journal = Journal of Bacteriology | volume = 169 | issue = 6 | pages = 2336–44 | date = June 1987 | pmid = 3584058 | pmc = 212055 | doi = 10.1128/jb.169.6.2336-2344.1987 | doi-access = free }}</ref> Both the ''virC'' and ''virD'' operons can be repressed by a chromosomally encoded protein known as Ros.<ref name="Cooley1991">{{cite journal | vauthors = Cooley MB, D'Souza MR, Kado CI | title = The virC and virD operons of the Agrobacterium Ti plasmid are regulated by the ros chromosomal gene: analysis of the cloned ros gene | journal = Journal of Bacteriology | volume = 173 | issue = 8 | pages = 2608–16 | date = April 1991 | pmid = 2013576 | pmc = 207827 | doi = 10.1128/jb.173.8.2608-2616.1991 | doi-access = free }}</ref><ref name="DSouza1993">{{cite journal | vauthors = D'Souza-Ault MR, Cooley MB, Kado CI | title = Analysis of the Ros repressor of Agrobacterium virC and virD operons: molecular intercommunication between plasmid and chromosomal genes | journal = Journal of Bacteriology | volume = 175 | issue = 11 | pages = 3486–90 | date = June 1993 | pmid = 8501053 | pmc = 204748 | doi = 10.1128/jb.175.11.3486-3490.1993 | doi-access = free }}</ref> Ros binds to a region of DNA that overlaps with the binding site of the VirG regulator, and therefore competes with VirG to control their expression levels.<ref name="Cooley1991" /><ref name="DSouza1993" /> Functionally, VirC1 and VirC2 promote the assembly of a [[Biology:Relaxosome|relaxosome]] complex during the conjugative transfer of T-DNA from the bacteria to the host plant cell.<ref name="Atmakuri2007" /> This is an energy-dependent process mediated via their NTPase activity, and occurs as they bind to a region of DNA known as ''overdrive''.<ref name="Atmakuri2007" /> As a result, they act to increase the amount of T-DNA strands produced. Following the production of the DNA strand to be transferred (transfer strand, T-strand), the VirC proteins can also help to direct the transfer strand to the transfer apparatus.<ref name="Atmakuri2007" />

The ''virD'' operon encodes for 4 proteins: VirD1-D4.<ref>{{cite journal | vauthors = Porter SG, Yanofsky MF, Nester EW | title = Molecular characterization of the virD operon from Agrobacterium tumefaciens | journal = Nucleic Acids Research | volume = 15 | issue = 18 | pages = 7503–17 | date = September 1987 | pmid = 3658701 | pmc = 306264 | doi = 10.1093/nar/15.18.7503 }}</ref> VirD1 and VirD2 are involved in the processing of T-DNA during conjugation to produce the T-strand; this is the single-stranded DNA molecule that is transported to the host plant cell (see transfer apparatus below).<ref name="Ghai1989">{{cite journal | vauthors = Ghai J, Das A | title = The virD operon of Agrobacterium tumefaciens Ti plasmid encodes a DNA-relaxing enzyme | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 86 | issue = 9 | pages = 3109–13 | date = May 1989 | pmid = 2541431 | pmc = 287074 | doi = 10.1073/pnas.86.9.3109 | doi-access = free | bibcode = 1989PNAS...86.3109G }}</ref> During the processing, VirD1 will act as a [[Biology:Topoisomerase|topoisomerase]] to unwind the DNA strands.<ref name="Ghai1989" /> VirD2, a [[Biology:Relaxase|relaxase]], will then nick one of the DNA strands and remain bound to the DNA as it is transferred to the recipient cell.<ref name="Zechner2012" /><ref>{{cite journal | vauthors = Yanofsky MF, Porter SG, Young C, Albright LM, Gordon MP, Nester EW | title = The virD operon of Agrobacterium tumefaciens encodes a site-specific endonuclease | journal = Cell | volume = 47 | issue = 3 | pages = 471–7 | date = November 1986 | pmid = 3021341 | doi = 10.1016/0092-8674(86)90604-5 | s2cid = 40721668 }}</ref> Within the recipient cell, VirD2 will also work together with VirE2 to direct the transferred DNA to the recipient cell's nucleus. There are suggestions that VirD2 may be phosphorylated and dephosphorylated by different proteins, affecting its ability to deliver DNA.<ref>{{cite journal | vauthors = Tao Y, Rao PK, Bhattacharjee S, Gelvin SB | title = Expression of plant protein phosphatase 2C interferes with nuclear import of the Agrobacterium T-complex protein VirD2 | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 101 | issue = 14 | pages = 5164–9 | date = April 2004 | pmid = 15047887 | pmc = 387391 | doi = 10.1073/pnas.0300084101 | doi-access = free | bibcode = 2004PNAS..101.5164T }}</ref> Conversely, little is known about VirD3, and mutational analyses have not provided any support for its role in the virulence of ''Agrobacterium''.<ref>{{cite journal | vauthors = Vogel AM, Das A | title = The Agrobacterium tumefaciens virD3 gene is not essential for tumorigenicity on plants | journal = Journal of Bacteriology | volume = 174 | issue = 15 | pages = 5161–4 | date = August 1992 | pmid = 1629176 | pmc = 206339 | doi = 10.1128/jb.174.15.5161-5164.1992 | doi-access = free }}</ref> Finally, VirD4 is a crucial part of the conjugation process, serving as a coupling factor that recognizes and transfers the T-strand to the transport channel.<ref>{{cite journal | vauthors = Kumar RB, Das A | title = Polar location and functional domains of the Agrobacterium tumefaciens DNA transfer protein VirD4 | journal = Molecular Microbiology | volume = 43 | issue = 6 | pages = 1523–32 | date = March 2002 | pmid = 11952902 | doi = 10.1046/j.1365-2958.2002.02829.x | s2cid = 43167663 }}</ref>

The ''virE'' operon encodes for 2 proteins: VirE1 and VirE2.<ref>{{cite journal | vauthors = Winans SC, Allenza P, Stachel SE, McBride KE, Nester EW | title = Characterization of the virE operon of the Agrobacterium Ti plasmid pTiA6 | journal = Nucleic Acids Research | volume = 15 | issue = 2 | pages = 825–37 | date = January 1987 | pmid = 3547330 | pmc = 340470 | doi = 10.1093/nar/15.2.825 }}</ref> VirE2 is an effector protein translocated together with the T-strand into host plant cells. There, it binds to the T-strand to direct its delivery to the [[Biology:Cell nucleus|nucleus]] of the host plant cell.<ref name="Galvin2012">{{cite journal | vauthors = Gelvin SB | title = Traversing the Cell: Agrobacterium T-DNA's Journey to the Host Genome | journal = Frontiers in Plant Science | volume = 3 | pages = 52 | date = 2012 | pmid = 22645590 | pmc = 3355731 | doi = 10.3389/fpls.2012.00052 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Das A | title = Agrobacterium tumefaciens virE operon encodes a single-stranded DNA-binding protein | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 85 | issue = 9 | pages = 2909–13 | date = May 1988 | pmid = 2452439 | pmc = 280112 | doi = 10.1073/pnas.85.9.2909 | doi-access = free | bibcode = 1988PNAS...85.2909D }}</ref> Part of this activity involves the presence of [[Biology:Nuclear localization sequence|nuclear localization sequence]]s within the protein, which marks the protein and the associated DNA for entry into the nucleus. It also protects the T-strand from [[Biology:Nuclease|nuclease]] attack.<ref name=Schrammeijer>{{cite journal | vauthors = Schrammeijer B, Beijersbergen A, Idler KB, Melchers LS, Thompson DV, Hooykaas PJ | title = Sequence analysis of the vir-region from Agrobacterium tumefaciens octopine Ti plasmid pTi15955 | journal = Journal of Experimental Botany | volume = 51 | issue = 347 | pages = 1167–9 | date = June 2000 | pmid = 10948245 | doi = 10.1093/jexbot/51.347.1167 | doi-access = free }}</ref> There is some speculation regarding the role of VirE2 as a protein channel, allowing DNA to move through the plant cytoplasmic membrane.<ref>{{cite journal | vauthors = Dumas F, Duckely M, Pelczar P, Van Gelder P, Hohn B | title = An Agrobacterium VirE2 channel for transferred-DNA transport into plant cells | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 98 | issue = 2 | pages = 485–90 | date = January 2001 | pmid = 11149937 | pmc = 14613 | doi = 10.1073/pnas.011477898 | doi-access = free | bibcode = 2001PNAS...98..485D }}</ref> On the other hand, VirE1 may be involved in promoting the transfer of the VirE2 protein into the host plant cell.<ref>{{cite journal | vauthors = Sundberg C, Meek L, Carroll K, Das A, Ream W | title = VirE1 protein mediates export of the single-stranded DNA-binding protein VirE2 from Agrobacterium tumefaciens into plant cells | journal = Journal of Bacteriology | volume = 178 | issue = 4 | pages = 1207–12 | date = February 1996 | pmid = 8576060 | pmc = 177787 | doi = 10.1128/jb.178.4.1207-1212.1996 | doi-access = free }}</ref> It binds to the ssDNA-binding domain of VirE2, therefore preventing the VirE2 protein from prematurely binding to the T-strand within the bacterial cell.<ref>{{cite journal | vauthors = Sundberg CD, Ream W | title = The Agrobacterium tumefaciens chaperone-like protein, VirE1, interacts with VirE2 at domains required for single-stranded DNA binding and cooperative interaction | journal = Journal of Bacteriology | volume = 181 | issue = 21 | pages = 6850–5 | date = November 1999 | pmid = 10542192 | pmc = 94155 | doi = 10.1128/JB.181.21.6850-6855.1999 }}</ref>

''virF'' is a host specificity factor found in some but not all types of Ti plasmids; for example, octopine-type Ti plasmids possess ''virF'' but nopaline-types do not.<ref name="Jarchow1991">{{cite journal | vauthors = Jarchow E, Grimsley NH, Hohn B | title = virF, the host-range-determining virulence gene of Agrobacterium tumefaciens, affects T-DNA transfer to Zea mays | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 88 | issue = 23 | pages = 10426–30 | date = December 1991 | pmid = 11607242 | pmc = 52941 | doi = 10.1073/pnas.88.23.10426 | doi-access = free | bibcode = 1991PNAS...8810426J }}</ref><ref name="Melchers1990">{{cite journal | vauthors = Melchers LS, Maroney MJ, den Dulk-Ras A, Thompson DV, van Vuuren HA, Schilperoort RA, Hooykaas PJ | title = Octopine and nopaline strains of Agrobacterium tumefaciens differ in virulence; molecular characterization of the virF locus | journal = Plant Molecular Biology | volume = 14 | issue = 2 | pages = 249–59 | date = February 1990 | pmid = 2101693 | doi = 10.1007/BF00018565 | s2cid = 8736045 }}</ref> The ability of ''A. tumefaciens'' to induce crown gall tumours in certain plant species but not others has been attributed to the presence or absence of this ''virF'' gene.<ref name="Jarchow1991" /><ref name="Melchers1990" />

The ''virH'' operon encodes for 2 proteins: VirH1 and VirH2.<ref name="Kalogeraki1999">{{cite journal | vauthors = Kalogeraki VS, Zhu J, Eberhard A, Madsen EL, Winans SC | title = The phenolic vir gene inducer ferulic acid is O-demethylated by the VirH2 protein of an Agrobacterium tumefaciens Ti plasmid | journal = Molecular Microbiology | volume = 34 | issue = 3 | pages = 512–22 | date = November 1999 | pmid = 10564493 | doi = 10.1046/j.1365-2958.1999.01617.x | doi-access = free }}</ref> A [[Biology:Bioinformatics|bioinformatics]] study of the amino acid sequences of the VirH protein showed similarities between them and a superfamily of proteins known as [[Biology:Cytochrome P450|cytochrome P450]] enzymes.<ref>{{cite journal | vauthors = Kanemoto RH, Powell AT, Akiyoshi DE, Regier DA, Kerstetter RA, Nester EW, Hawes MC, Gordon MP | display-authors = 6 | title = Nucleotide sequence and analysis of the plant-inducible locus pinF from Agrobacterium tumefaciens | journal = Journal of Bacteriology | volume = 171 | issue = 5 | pages = 2506–12 | date = May 1989 | pmid = 2708311 | pmc = 209927 | doi = 10.1128/jb.171.5.2506-2512.1989 | doi-access = free }}</ref> VirH2 was then discovered to metabolize certain phenolic compounds detected by VirA.<ref name="Kalogeraki1999" />

===Transfer DNA (T-DNA)===
The T-DNA of ''Agrobacterium'' is approximately 15-20 kbp in length and will become integrated into the host plant genome upon its transfer via a process known as [[Biology:Genetic recombination|recombination]]. This process utilizes preexisting gaps in the host plant cell's genome to allow the T-DNA to pair with short sequences in the genome, priming the process of [[Biology:Ligation (molecular biology)|DNA ligation]], where the T-DNA is permanently joint to the plant genome.<ref name="Galvin2012" /> The T-DNA region is flanked at both ends by 24bp sequences.

Within the host plant cell's genome, the T-DNA of ''Agrobacterium'' is expressed to produce two main groups of proteins.<ref name="Gordon2014" /> One group is responsible for the production of plant growth hormones. As these hormones are produced, there will be an increase in the rate of cell division and therefore the formation of crown gall tumors.<ref name="Zhu2000" /> The second group of proteins are responsible for driving the synthesis of opines in the host plant cells. The specific opines produced depends on the type of the Ti plasmid but not on the plant host. These opines cannot be utilized by the plant host, and will instead be exported out of the plant cell where it can be taken up by the ''Agrobacterium'' cells. The bacteria possess genes in other regions of the Ti plasmid that allows the catabolism of opines.<ref name="Gordon2014" />

===Transfer apparatus===
Transfer apparatuses encoded within the Ti plasmid have to achieve two objectives: allow the conjugative transfer of the Ti plasmid between bacteria and allow the delivery of the T-DNA and certain effector proteins into host plant cells. These are achieved by the Tra/Trb system and the VirB/VirD4 system respectively, which are members of the [[Biology:Bacterial secretion system|type IV secretion system]] (T4SS).<ref name="Zhu2000">{{cite journal | vauthors = Zhu J, Oger PM, Schrammeijer B, Hooykaas PJ, Farrand SK, Winans SC | title = The bases of crown gall tumorigenesis | journal = Journal of Bacteriology | volume = 182 | issue = 14 | pages = 3885–95 | date = July 2000 | pmid = 10869063 | pmc = 94570 | doi = 10.1128/jb.182.14.3885-3895.2000 | doi-access = free }}</ref>

For the Ti plasmid and T-DNA to be transferred via conjugation, they must first be processed by different proteins, such as the relaxase enzyme (TraA/VirD2) and the DNA transfer and replication (Dtr) proteins. Together, these proteins will recognize and bind to a region known as the [[Biology:Origin of transfer|origin of transfer]] (''oriT'') in the Ti plasmid to form the relaxosome complex. For the T-DNA, a nick will be created at the T-DNA's border sequence, and the nicked T-strand will be transported to the cell membrane, where the rest of the transfer machinery is present.<ref name="Zechner2012">{{cite journal | vauthors = Zechner EL, Lang S, Schildbach JF | title = Assembly and mechanisms of bacterial type IV secretion machines | journal = Philosophical Transactions of the Royal Society of London. Series B, Biological Sciences | volume = 367 | issue = 1592 | pages = 1073–87 | date = April 2012 | pmid = 22411979 | pmc = 3297438 | doi = 10.1098/rstb.2011.0207 | doi-access = free }}</ref>

Within the VirB/VirD4 system, the VirD2 relaxase is aided by the accessory factors VirD1, VirC1 and VirC2 while it processes the DNA substrate.<ref>{{cite journal | vauthors = De Vos G, Zambryski P | title = Expression of Agrobacterium nopaline-specific VirD1, VirD2, and VirC1 proteins and their requirement for T-strand production in E. coli | journal = Molecular Plant-Microbe Interactions | volume = 2 | issue = 2 | pages = 43–52 | date = 1989 | pmid = 2520160 | doi = 10.1094/mpmi-2-043 }}</ref> Furthermore, the VirD2 relaxase and the VirC proteins will contribute to the delivery of the DNA strand to the VirD4 receptor at the cell membrane.<ref name="Atmakuri2007">{{cite journal | vauthors = Atmakuri K, Cascales E, Burton OT, Banta LM, Christie PJ | title = Agrobacterium ParA/MinD-like VirC1 spatially coordinates early conjugative DNA transfer reactions | journal = The EMBO Journal | volume = 26 | issue = 10 | pages = 2540–51 | date = May 2007 | pmid = 17505518 | pmc = 1868908 | doi = 10.1038/sj.emboj.7601696 }}</ref> This receptor is an essential component of T4SSs, and is thought to energize and mediate the transfer of the DNA into the translocation channel between two cells.<ref>{{cite journal | vauthors = Gomis-Rüth FX, Solà M, de la Cruz F, Coll M | title = Coupling factors in macromolecular type-IV secretion machineries | journal = Current Pharmaceutical Design | volume = 10 | issue = 13 | pages = 1551–65 | date = 2004 | pmid = 15134575 | doi = 10.2174/1381612043384817 }}</ref> The table below summarizes the proteins encoded in the ''virB'' operon that makes up the translocation channel of the VirB/VirD4 system.<ref name="Gordon2014" />

{| class="wikitable"
|-
! Protein(s)
! Function
|-
| VirB4, VirB11
| ATPases that provide the energy for DNA transfer<ref name="christie2005">{{cite journal | vauthors = Christie PJ, Atmakuri K, Krishnamoorthy V, Jakubowski S, Cascales E | title = Biogenesis, architecture, and function of bacterial type IV secretion systems | journal = Annual Review of Microbiology | volume = 59 | pages = 451–85 | date = 2005 | pmid = 16153176 | pmc = 3872966 | doi = 10.1146/annurev.micro.58.030603.123630 }}</ref><ref>{{cite journal | vauthors = Peña A, Matilla I, Martín-Benito J, Valpuesta JM, Carrascosa JL, de la Cruz F, Cabezón E, Arechaga I | display-authors = 6 | title = The hexameric structure of a conjugative VirB4 protein ATPase provides new insights for a functional and phylogenetic relationship with DNA translocases | journal = The Journal of Biological Chemistry | volume = 287 | issue = 47 | pages = 39925–32 | date = November 2012 | pmid = 23035111 | pmc = 3501061 | doi = 10.1074/jbc.M112.413849 | doi-access = free }}</ref>
|-
| VirB3, VirB6, VirB8
| Subunits of a putative inner membrane [[Biology:Translocase|translocase]]<ref name="christie2005" /><ref>{{cite journal | vauthors = Mossey P, Hudacek A, Das A | title = Agrobacterium tumefaciens type IV secretion protein VirB3 is an inner membrane protein and requires VirB4, VirB7, and VirB8 for stabilization | journal = Journal of Bacteriology | volume = 192 | issue = 11 | pages = 2830–8 | date = June 2010 | pmid = 20348257 | pmc = 2876495 | doi = 10.1128/JB.01331-09 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Jakubowski SJ, Krishnamoorthy V, Cascales E, Christie PJ | title = Agrobacterium tumefaciens VirB6 domains direct the ordered export of a DNA substrate through a type IV secretion System | journal = Journal of Molecular Biology | volume = 341 | issue = 4 | pages = 961–77 | date = August 2004 | pmid = 15328612 | pmc = 3918220 | doi = 10.1016/j.jmb.2004.06.052 }}</ref>
|-
| VirB7, VirB9, VirB10
| Forms a core complex that stabilizes the channel subunits<ref name="christie2005" /><ref>{{cite journal | vauthors = Chandran V, Fronzes R, Duquerroy S, Cronin N, Navaza J, Waksman G | title = Structure of the outer membrane complex of a type IV secretion system | journal = Nature | volume = 462 | issue = 7276 | pages = 1011–5 | date = December 2009 | pmid = 19946264 | pmc = 2797999 | doi = 10.1038/nature08588 | bibcode = 2009Natur.462.1011C }}</ref> 
|-
| VirB2
| The major [[Biology:Pilin|pilin]] subunit of the conjugative [[Biology:Pilus|pilus]]<ref name="christie2005" />
|-
| VirB1, VirB5
| Minor components of the conjugative pilus<ref>{{cite journal | vauthors = Zupan J, Hackworth CA, Aguilar J, Ward D, Zambryski P | title = VirB1* promotes T-pilus formation in the vir-Type IV secretion system of Agrobacterium tumefaciens | journal = Journal of Bacteriology | volume = 189 | issue = 18 | pages = 6551–63 | date = September 2007 | pmid = 17631630 | pmc = 2045169 | doi = 10.1128/JB.00480-07 | doi-access = free }}</ref><ref>{{cite journal | vauthors = Schmidt-Eisenlohr H, Domke N, Angerer C, Wanner G, Zambryski PC, Baron C | title = Vir proteins stabilize VirB5 and mediate its association with the T pilus of Agrobacterium tumefaciens | journal = Journal of Bacteriology | volume = 181 | issue = 24 | pages = 7485–92 | date = December 1999 | pmid = 10601205 | pmc = 94205 | doi = 10.1128/JB.181.24.7485-7492.1999 | doi-access = free }}</ref>
|}

==Uses in bioengineering==
The ability of ''Agrobacterium'' to deliver DNA into plant cells opened new doors for plant genome engineering, allowing the production of genetically modified plants (transgenic plants).<ref>{{cite journal| vauthors = Hernalsteens JP, Van Vliet F, De Beuckeleer M, Depicker A, Engler G, Lemmers M, Holsters M, Van Montagu M, Schell J | s2cid=4333703| title=The ''Agrobacterium tumefaciens'' Ti plasmid as a host vector system for introducing foreign DNA in plant cells|date=1980|journal=Nature|volume=287 |issue=5783 |pages=654–656| doi=10.1038/287654a0| bibcode=1980Natur.287..654H}}</ref> Proteins involved in mediating the transfer of T-DNA will first recognize the border sequences of the T-DNA region. Therefore, it is possible for scientists to use T-DNA border sequences to flank any desired sequence of interest - such a product can then be inserted into a plasmid and introduced into ''Agrobacterium'' cells.<ref>{{cite journal | vauthors = Gelvin SB | title = Agrobacterium-mediated plant transformation: the biology behind the "gene-jockeying" tool | journal = Microbiology and Molecular Biology Reviews | volume = 67 | issue = 1 | pages = 16–37, table of contents | date = March 2003 | pmid = 12626681 | pmc = 150518 | doi = 10.1128/mmbr.67.1.16-37.2003 | doi-access = free }}</ref> There, the border sequences will be recognized by the transfer apparatus of ''A. tumefaciens'' and delivered in a standard manner into the target plant cell.<ref name="Gordon2014" /> Moreover, by leaving behind only the border sequences of the T-DNA, the resulting product will edit the plant genome without causing any tumours in plants.<ref>{{cite journal | vauthors = Zambryski P, Joos H, Genetello C, Leemans J, Montagu MV, Schell J | title = Ti plasmid vector for the introduction of DNA into plant cells without alteration of their normal regeneration capacity | journal = The EMBO Journal | volume = 2 | issue = 12 | pages = 2143–50 | date = 1983 | pmid = 16453482 | doi = 10.1002/j.1460-2075.1983.tb01715.x | pmc = 555426 }}</ref> This method has been used to modify several crop plants, including rice,<ref>{{cite journal|vauthors=Chan MT, Lee TM, Chang HH | title=Transformation of indica rice (''Oryza sativa'' L.) mediated by ''Agrobacterium tumefaciens|date=1992|journal=Plant and Cell Physiology|volume=33 |issue=5 |pages=577–583 |doi=10.1093/oxfordjournals.pcp.a078292}}</ref> barley<ref>{{cite journal| vauthors = Tingay S, McElroy D, Kalla R, Fieg S, Wang M, Thornton S, Brettell R | title=''Agrobacterium tumefaciens''-mediated barley transformation|date=1997|journal=The Plant Journal|volume=11 |issue=6 |pages=1369–1376 |doi=10.1046/j.1365-313X.1997.11061369.x|doi-access=free}}</ref> and wheat.<ref>{{cite journal | vauthors = Cheng M, Fry JE, Pang S, Zhou H, Hironaka CM, Duncan DR, Conner TW, Wan Y | display-authors = 6 | title = Genetic Transformation of Wheat Mediated by Agrobacterium tumefaciens | journal = Plant Physiology | volume = 115 | issue = 3 | pages = 971–980 | date = November 1997 | pmid = 12223854 | pmc = 158560 | doi = 10.1104/pp.115.3.971 | doi-access = free }}</ref> Further work have since extended the targets of ''A. tumefaciens'' to include fungi and human cell lines.<ref>{{cite book | vauthors = Tzfira T, Citovsky V |title=''Agrobacterium'': from biology to biotechnology |date=2007 |publisher=Springer Science & Business Media}}</ref><ref>{{cite journal | vauthors = Kunik T, Tzfira T, Kapulnik Y, Gafni Y, Dingwall C, Citovsky V | title = Genetic transformation of HeLa cells by Agrobacterium | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 98 | issue = 4 | pages = 1871–6 | date = February 2001 | pmid = 11172043 | pmc = 29349 | doi = 10.1073/pnas.98.4.1871 | doi-access = free | bibcode = 2001PNAS...98.1871K }}</ref>

== Similar plasmids ==
<!-- Something about RepABC incompatibility groups... use doi:10.1007/978-3-540-85467-8_6-->

=== Root-inducing (Ri) plasmid ===

=== Symbiotic (sym) plasmids of Rhizobia ===

== See also ==
* Mary-Dell Chilton
* Jeff Schell
* Marc Van Montagu

== References ==
{{reflist}}

== External links ==
* [https://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=genome&cmd=Retrieve&dopt=Overview&list_uids=15251 Agrobacterium tumefaciens plasmid pTi-SAKURA, complete sequence]
* [http://micro.cornell.edu/research/winans-lab/ti-plasmid-genetic-map Ti Plasmid Genetic Map]
* [https://www.rhs.org.uk/advice/profile?pid=141 Crown gall disease]

{{Self-replicating organic structures}}

{{DEFAULTSORT:Ti Plasmid}}
[[Category:Plasmids]]

{{Sourceattribution|Ti plasmid}}