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Summary of patents on modified co-integrated vectors and methods

Several entities own inventions built on the basic components of co-integrated vectors and the basic methods to assemble them. Three different inventions are referred to in this section:

  • Stable co-integrated vector. The invention of Schilperoort and Hille in the United States and in Europe discloses a way to combine in E. coli a vector molecule (pR772) having a modified T-region with a Ti plasmid (pTiB6). After the recombination of the molecules, the co-integrated plasmid pAL969 is transferred to an Agrobacterium strain lacking a native Ti-plasmid. The co-integrated vector and its derivatives are stable in both E.coli and Agrobacterium and are used for transformation of dicot plants.The very narrow scope of this patent will not present undue difficulties for most researchers. Quite simply, by using different plasmids than those claimed, infringement of this patent can likely be avoided.
  • Site-specific co-integrated vector. The granted United States patent and the European application assigned to Mogen, now called Syngenta Mogen B.V., discloses an alternate way to attain homologous recombination in order to form a co-integrated vector. The recombination process is triggered by the action of Cre (causing recombination) recombinase on recombination sites (i.e. lox P) present in both a small vector and a disarmed (tumor-genes free) Ti plasmid. The following diagram illustrates this method.
    In conclusion, the most limiting factor in this patent is the presence and use of the Cre recombinase system to form a stable co-integrated plasmid within an Agrobacteriumstrain.
  • Co-transformation with two different T-DNAs. Japan Tobacco has one United States granted patent and a European application directed to this subject matter. This invention discloses an Agrobacterium strain containing a first T-DNA, which has a selectable marker, e.g. a drug resistance gene, and a second T-DNA, which has a desired gene or a cloning site and is part of a co-integrated vector. The first T-DNA may or may not be part of the same vector as the second T-DNA. Although when both are in the same co-integrated vector, they are distant enough to be inherited independently.Thus, an Agrobacterium strain having two different T-DNAs as described above is very likely to fall within the scope of the claimed invention in the United States patent. Note, however, that the second vector used in this invention must be a co-integrated vector.

Table of patents on modified co-integrated vectors

The following table presents an overview of six patents and patent applications that claim co-integrated vectors having variations on their basic elements.

This analysis is limited by being based only upon the published specification and claims of the issued patents and patent applications.

Assigned to Schilperoort & Hille
Issued Patents Overview
US 4693976

Method for incorporating foreign DNA into a dicot genome via A. tumefaciens using a stable co-integrated plasmid. The co-integrated plasmid, composed of a Ti-plasmid (pTiB6) and a broad host range plasmid (R772), contains foreign DNA in the T-region of the Ti component of the co-integrated plasmid. The insertion of the new genes into the T-region of the R::Ti co-integrate is achieved by homologous recombination of an engineered E.coli vector with a modified T-region and the R::Ti co-integrate.

EP 120515 B1 
Process for incorporating foreign DNA into a dicot genome by infecting plant protoplasts with A. tumefaciens containing a plasmid derived from the co-integrated plasmid pAL969. Such plasmid contains only foreign DNA between the 23 base pairs of the wild-type T-region from the Ti plasmid pTiB6.
Assigned to Syngenta Mogen B.V.
Issued Patents Overview
US 5635381

Agrobacterium strains having a site-specific recombinase (Cre) capable of causing recombination between two different recombination sites (i.e. loxP site) present in the same strains. The site-specific recombination event between two separated DNA molecules each carrying a recombination site results in the formation of a site-specific co-integrated plasmid.

EP 628082 B1
This recently granted patent (former application EP 628082 A1 ) is directed to an Agrobacterium strain having a gene encoding a recombinase and a sequence capable of controlling its expression. The recombinase causes site-specific recombination between two recombination sites. The scope of the claims as granted is the same as the claims filed in the EP application.
Assigned to Japan Tobacco Inc.
Issued Patents Overview Patent Applications Overview
US 5731179

Method for co-transforming plant cells with two T-DNAs via Agrobacterium where the first T-DNA contains a selectable marker, and the second T-DNA has a gene of interest. Either both T-DNAs are located in the same hybrid vector or at least the second T-DNA is in a hybrid vector, which is the product of homologous recombination between an acceptor vector and an intermediate vector.

 EP 687 730 A1

Method for co-transforming plant cells with a first T-DNA containing a drug resistance gene and a second T-DNA containing a desired gene and contained in a hybrid vector prepared by homologous recombination between an acceptor vector and an intermediate vector. A hybrid vector with a first T-DNA as described and a second T-DNA with a cloning site is further claimed.