Ti-Plasmid

Tumor -inducing plasmids (english tumor inducing plasmid ) are plasmids that often, but not always, for the genetic makeup of the bacteria Agrobacterium tumefaciens and Agrobacterium rhizogenes are. The genes of the Ti plasmid, enable Agrobacterium DNA transfer into plant cells and to manipulate genetically. They trigger tumor-like growths, and thus plant diseases.

Genes of Ti plasmids

The Ti plasmid is 200 kb large. The following groups of important genes are located on the plasmid:

  • Transfer or tra genes
  • Virulence or vir genes (both essential for DNA transfer to the plant cell )
  • Genes for Opinkatabolismus ( necessary for the degradation of opines by the bacterium )

And transmitted to the plant cell or T-DNA transfer with

  • Tumorigenesis or onc genes (required for tumor induction in the plant cell, for example, TMS1, TMS2 and TMR), as well as
  • Opine synthesis genes or OPS ( necessary for the synthesis of opines by the plant cell, for example for the nos nopaline synthesis OCS for octopine synthesis etc.)

The T- DNA is flanked by imperfect tandem repeats of 25 bases: "left border" (LB ) and "right border" (RB ) indicate the Vir proteins beginning and end of the T- DNA. These cis-elements are essential for the transfer of the T-DNA.

The plasmid also has an origin of replication (ori or origin of replication ) and other regions. The genes for the identification of plant cells and the attachment of these on the other hand are located in the bacterial genome.

Mechanism of gene activation and transmission

Acetosyringone and other phenolic compounds that emerge from wounded plant parts, select the one located in the outer bacterial cell membrane sensor kinase product of the virA gene. This phosphorylated and activates the VirG protein. VirG is a response regulator, and that an activator of other vir genes whose transcription rate hereby increases by a multiple. Among the now transcribed genes include virD, Vire and vir. VirD is an endonuclease. The protein causes the right repeat flanking the T-DNA, a single-strand break. This begins the synthesis of a complementary strand; the single strand is displaced thereby. This mechanism is similar to the rolling circle replication of plasmids, only that arises here is a straight single strand with a given end. Vire binds and stabilizes the single-stranded T- DNA and protects it from degradation. Vir B is located in the bacterial cell membrane and allows direct contact with the plant cell the transfer of the T-DNA. In addition, the adhesion proteins chvA, chvB and PSCA on the T- DNA transfer are involved. The mechanism of DNA transfer is similar to bacterial conjugation. VirD the protein remains in the T-DNA bound and enters this into the plant cell. The transferred T- DNA is about 20 kb in size.

Integration of T- DNA

The protein has a vire Kernlokalisatiossequenz and transported from their entry into the plant cell together with the T-DNA -coated along the cytoskeleton in the plant cell nucleus. The integration into the plant genome occurs rather non-specific, but is favored by certain tandem repeats.

Effect of T-DNA

The onc genes include, among others, the genes for the Tryptophanmonooxygenase (Gen IAAM, with iaa stands for indole -3 -acetic acid, the biologically active plant hormone auxin ), indoleacetamide ( IAAH ) and the isopentenyl transferase ( itpZ ). These enzymes disrupt the hormonal balance of the plant by an additional synthesis of the plant hormones auxin and cytokinin is induced. The cells begin to divide; the growths known as " bile " arise. The induced by Ti plasmids tumors usually consist of little or undifferentiated callus. The by Ri plasmids of Agrobacterium rhizogenes -induced proliferation of roots, however, provide at least partially differentiated root tissue dar. The ops genes cause the plant cell to produce nitrogen-rich opines. Since the genes for the catabolism (breakdown) of opines are not transferred, the bacteria is such an exclusive energy and nitrogen source. It is interesting that the genes derived from the bacterium of the t- DNA have a typical eukaryotic structure and in the plant cell are active.

Genetic application

The genetic modification of plants is comparatively difficult. For this reason, the natural gene transfer system has been studied extensively and used in plant cells. In the transformation of plants, Ti plasmids are used, in which the genes have been substituted for the tumor formation by the desired genes. The addition of nematodes causes the violation of the plants at the cellular level and paves the way for infection by Agrobacterium.

The system of the plant transformation using Agrobacterium is successfully applied to the study of plant genetics. In addition, it is used for production of commercial genetically modified plants. Examples are

  • Bt maize ( " GM maize " )
  • Flavr Savr tomato ( " Flavr Savr tomato" )
  • Herbicide resistant corn

And others.

Binary vectors

To keep the amount of foreign genes that are transferred to the plant, as low as possible, it uses the system of binary vectors. Binary vectors are the most widely used system in A. tumefaciens -mediated transfer of genes to plants. With this system in the bacterium are two plasmids. A disarmed Ti - plasmid carrying the vir region, the T-DNA region but was removed; called "helper Ti - plasmid". In addition, a plasmid containing the ' gene of interest ' carries a T-DNA region that is to be transmitted. In the infection of plants with A. tumefaciens carrying the two plasmids, the helper Ti - plasmid triggers the association of the bacteria to the plant cell. Subsequently, the T- DNA of the plasmid from the second bacterium is transferred into the plant genome.

Credentials

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