Molecular farming

Molecular farming:- It describes the production of recombinant proteins and other secondary metabolites in plants. This technology depends on a genetic transformation of plants.

The strategies of plant transformation:- Molecular farming depending on the production of transgenic plants. It has been operated by two general methods:

1. Stable or permanent expression systems:-

a. Stable nuclear transformation:- 

> It refers to the integration of foreign genes into the nuclear genome of plants, which results in the change of genetic structures and consequent expression of transgenes after integration with the host genomes. 

> The largest amount of recombinant proteins has been produced by one of the most common methods of stable nuclear transformation. 

> A method exploited for aggregating proteins in dried beans of maize culminates in a long-term storage in the beans at the room temperature without decomposition of proteins. 

> In addition, it has a considerable potential for producing crops like cereals that actually grow everywhere. 

> Limitations:- 

i. A long production cycle for some crops.

ii. Potential collisions of some crops with natural species or food products.

b. Stable plastid transformation:- 

> Numerous advantages:-

i. Preventing transgene escape through amphimixis, because plastids are inherited through the maternal tissue in the majority of species.

ii. Absence of chloroplasts in pollen and consequent improbability of their transfer, which reduces environmental concerns. 

> The transformed transgenic plants with homoplasmic chloroplasts (all chloroplasts carry transgenes) were selected after several generations of plant regeneration from bombarded leaf explants. 

> Selection was conducted on a medium containing spectinomycin or combined with streptomycin. 

> The researchers have already extracted a human pharmaceutical protein, more than 3% to 6%, from the total soluble proteins in the chloroplasts of tobacco. 

> Recently, Oey reported a very high level (70% of an entire soluble protein) for a protein antibiotic with the chloroplast system, which, till today, has been the highest concentration of recombinant proteins. 

> Functional limitation:- 

i. Chloroplast transformation only in tobacco is practically possible, but unfortunately this plant is inedible and full of poisonous alkaloids. 

ii. Long lasting storage in refrigerators will bring about changes in protein stability.

c. Plant cell suspension culture:- 

> This method involves the removal of cell walls and gene transfer to the obtained protoplasts and suspension culture. 

> The purification system and its downstream processing are cheaper and easier. 

> In addition, the use of suspension culture can decrease heterogeneity in proteins and sugar (N-glycans) regarding the uniformity of the type and size of cells. 

> Furthermore, as a fast system there is no need for the production of transgenic plants. 

> Some samples of plant suspension cultures can be used for producing biomedicines, including:-

i. Vaccines of Newcastle disease virus of chicks

ii. Recombinant human glucocerebrosidase for treating diabetes. 

> Though this method is cheaper, safer, and easier in comparison to the other methods of genetic manipulation, cell suspension has not yet been suggested as an optimal production choice of production in plant systems. 

> Limitation:- It is believed that the ultimate products and their usability are constrained by reducing the level of recombinant proteins during the stationary phase because of the enhanced proteolytic activity.

2. Temporary or transient expression systems:-

a. Agrobacterium transformation method:- 

> Infiltration of recombinant agrobacterium suspension into tobacco leaf tissue is achieved without stable gene transfer, which facilitates the transfer of T-DNA to a very high percentage of cells, where the transgenes are expressed at a high level without a stable transfer of genes. 

> Presently, this method has been very efficient for the production of clinical biomedicines with a fast expansion.

b. Viral infection methods:- The viral infection method depends on the capability of plant viruses, such as tobacco mosaic virus and X potato virus, which functions as a vector to convey foreign genes into plant genomes without fusing with the genome of that plant.

c. Magnifection system:- 

> Expression systems based on viral vectors and agrobacterium methods suffer from some constraints for the co-expression of two or more polypeptides required for the production of hetero-oligomeric proteins. 

> Thus, a new transient expression system known as MagnICON technology has been developed by Icon Genetics Company. 

> This method includes removing coat proteins (responsible for systemic movement) of non-competitive virus stains and systemic delivery of the derived viral vectors to all of the plants using agrobacterium as the medium of primary infection. 

> This method not only optimizes the infection but also significantly increases proliferation, and finally results in the co-expression of several polypeptides and the rise of functional protein production more than 100 times.

The advantages of utilizing transgenic plants as bioreactors:- Comparison of different expression systems reveals the advantages of plants in comparison with other expression systems as follows:

i. The healthiness of derived products (plants cannot be the host of human pathogens and bacterial toxins).

ii. Capability of post-translational processing (respecting the features of eukaryotic cells).

iii. The possibility of using breeding methods and sexual crosses to obtain active recombinant multi-chain proteins (therefore, there is the possibility of producing antibodies without application of a double transformation).

iv. Reducing the costs of production (plants can produce biological materials by the use of carbon dioxide, solar energy, and inorganic materials. Moreover, the scale of production can be manipulated regarding scalability).

v. Reducing the costs of storage and transportation of recombinant proteins (when they are produced in dry textures like grains).

vi. Removing the purification step (when the plant tissues containing recombinant protein are edible).