Methods of Gene Transfer in Plants
To add a desired trait to a crop, a foreign gene (transgene) encoding the trait must be inserted into plant cells, along with a “cassette” of additional genetic material. The cassette includes a DNA sequence called a “promoter,” which determines where and when the foreign gene is expressed in the host, and a “marker gene” that allows breeders to determine which plants contain the inserted gene by screening or selection. For example, marker genes may render plants resistant to antibiotics that are not used medically (e.g., agromycin, canamycin) or tolerant to certain herbicides.
Two methods are used to transfer foreign genes into plants. The first method involves the use of a plant pathogen called Agrobacterium tumefaciens, which causes crown gall disease in many species. This bacterium has a plasmid, or loop of non-chromosomal DNA, that contains tumor-inducing genes (T-DNA), along with additional genes that help the T-DNA integrate into the host genome. For genetic engineering purposes, Agrobacterium must first be “disarmed” so that it does not make the plant sick. This is done by removing most of the T-DNA while leaving the left and right border sequences, which integrate a foreign gene into the genome of cultured plant cells.
The second delivery method is a “gene gun,” which fires gold particles carrying the foreign DNA into plant cells. Some of these particles pass through the plant cell wall and enter the cell nucleus, where the transgene integrates itself into the plant chromosome. Because both methods of gene transfer are fairly random, one must screen for the plant cells that contain the foreign gene.