Intragenic vectors for gene transfer without foreign DNA

Abstract

The intragenic vector system involves identifying functional equivalents of vector components from the genome of a specific crop species (or related species to which it can be hybridised) and using these DNA sequences to assemble vectors for transformation of that plant species. This system offers an attractive alternative to current genetic engineering strategies where vectors are based on DNA sequences that usually originate from bacteria. The construction of intragenic vectors enables the well-defined genetic improvement of plants with all transferred DNA originating from within the gene pool already available to plant breeders. In this manner genes can be introgressed into elite cultivars in a single step without linkage drag and without the incorporation of foreign DNA. The resulting plants are non-transgenic, although they are derived using the tools of molecular biology and plant transformation. The use of intragenic vectors for the transfer of genes from within the gene pools of crops may help to alleviate some of the major public concerns over the deployment of GM crops in agriculture, notably the ethical issue associated with the transfer of DNA across wide taxonomic boundaries. This paper reviews the progress toward the development and use of intragenic vectors and the implications of their use for the genetic improvement of crops.

A Set of Modular Binary Vectors for Transformation of Cereals

“…Compared to biolistic techniques (Stoeger et al, 1999;Bhalla et al, 2006) Agrobacterium-mediated transformation offers several advantages (Tzfira and Citovsky, 2006), such as simpler integration patterns resulting in lower mutational consequences for the transgenic plant (Latham et al, 2006) and limited transgene silencing via cosuppression. In addition, the option for fine tuning the Agrobacterium-based transformation protocols renders more and more cereal species amenable for efficient genetic engineering (Shrawat and Loerz, 2006;Conner et al, 2007). These advantages prompted us to implement the GATEWAY cloning system and expression cassettes into a vector set for Agrobacteriummediated transformation, thereby providing the versatility to use the vector set for the transformation of a large panel of cereal species and genotypes.…”

Genetic transformation of apple (Malus x domestica) without use of a selectable marker gene

“…Essential components of the binary vector currently used are derived from bacterial systems, such as the T-DNA border regions and the DNA into which the gene of interest is cloned. To avoid such problem, Conner et al (2007) have developed the concept of intragenic vectors consisting of only plant-derived DNA fragments. They have developed this type of vector for tobacco.…”

“…Current sequencing of the apple genome will provide the information necessary to identify DNA fragments with functional equivalence of important vector components. Already, Conner et al (2007) have developed some T-DNA-like regions for apple. This type of intragenic vector and MDDT will allow production of "intragenic" (Nielsen 2003), "all native" (Rommens 2004), or "cisgenic" (Schouten et al 2006a, b) plants for highly targeted genetic improvement.…”

Citrus Transformation: Challenges and Prospects

“…Such genes could be perceived by the public as more "natural" and could potentially be less likely to be toxic or allergenic (although this would have to be tested experimentally on a case by case basis). Plants transformed this way do not appear to raise the fear and ethical concerns that the production of transgenic plants inspires (Conner et al, 2006;Rommens et al, 2007). However, this approach would rule out the use of most commonly used selectable and scorable marker genes, as well as the most commonly used promoters and termination sequences and the necessary T-DNA borders for Agrobacterium-mediated transformation.…”