Plastid transformation

Plastid transformation is a method in the genetic engineering of plants. Instead of the nuclear DNA, the DNA in the plant's plastids, usually the chloroplasts, is modified. The major advantage of this technology is that in many plant species plastid DNA is not inherited, which prevents gene flow from the genetically modified plant to other plants.

Transformation technology
The most common method to transform plastids is particle bombardment: Small gold or tungsten particles are coated with DNA and shot into young plant cells or plant embryos. Some genetic material will stay in the cells and transform them. The transformation efficiency is lower than in agrobacterial mediated transformation, which is also common in plant genetic engineering, but particle bombardment is especially suitable for plastid transformation.

In order to persist and be stably maintained in the cell, a plasmid DNA molecule must contain an origin of replication, which allows it to be replicated in the cell independently of the chromosome. Because transformation usually produces a mixture of rare transformed cells and abundant non-transformed cells, a method is needed to identify the cells that have acquired the plasmid. Plasmids used in transformation experiments will usually also contain a gene giving resistance to an antibiotic that the intended recipient strain of bacteria is sensitive to. Selection for cells able to grow on media containing this antibiotic can then select the cells that have acquired the plasmid by transformation, as cells lacking the plasmid will be unable to grow.

Biological containment and agricultural coexistence
Genetically modified plants must be safe for the environment and suitable for coexistence with conventional and organic crops. Towards such safety, a major hurdle is posed by the potential outcrossing of the transgene via pollen movement. Plastid transformation, which yields transplastomic plants in which the pollen does not contain the transgene, not only increases biosafety, but also facilitates the coexistence of genetically modified, conventional and organic agriculture. Therefore, developing such crops is a major goal of research projects such as Co-Extra and Transcontainer.

Transplastomic tobacco
However, plastid transformation is suitable only for certain crop species, and the reliability of this method has only been proven for tobacco. Led by Ralph Bock from the Max Planck Institute of Molecular Plant Physiology in Germany, researchers studied genetically modified tobacco in which the transgene was integrated in chloroplasts. Since past literature reported contradicting figures on the reliability of this process, the Co-Extra researchers analysed more than two million seedlings and found that less than 20 in 1,000,000 inherited the transgene. In the pollen of adult plants, the rate was even lower, remaining below 3 in 1,000,000. This reduction is due to the fact that some parts of the seedlings are lost during their development into mature plants.

Because tobacco has a strong tendency towards self-fertilisation, the reliability of transplastomic plants is assumed to be even higher under field conditions. Therefore, the researchers believe that only one in 100,000,000 GM tobacco plants actually would transmit the transgene via pollen. Such values are more than satisfactory to ensure coexistence. However, for GM crops used in the production of pharmaceuticals, or in other cases in which absolutely no outcrossing is permitted, the researchers recommend the combination of chloroplast transformation with other biological containment methods, such as cytoplasmic male sterility or transgene mitigation strategies.