Basic Concept of Biotechnology 23

  Plant Molecular Farming: A Promising Stratergy in Biotechnology    The improvement of agricultural crop plants relied largely on the conventional breeding programs to increase the productivity, alter the quality characteristics or impart resistance to biotic or abiotic stresses. However, with the advancement of molecular biology techniques it has become possible to introduce entirely new characteristics efficiently through insertion of the genes coding for the desired characteristics directly into the genome of the plant or animal. Also, these techniques have made possible to introduce non-native genes from varied sources, thus tailoring the plant or animal to produce entirely new products for innovative applications. A large number of recombinant proteins have been produced in plants over the last twenty years, demonstrating the ability of plants to compete with existing industrial production systems. The use of plants for producing recombinant proteins has been termed...

  Plants as Expression Systems Plants are modified to produce a wide range of heterologous proteins including pharmaceutical and industrial proteins, through recombinant DNA technology, often referred as plant molecular farming (Faye and Gomord, 2010; Ma and Wang, 2012; Obembe  et al ., 2011; Wilken and Nikolov, 2012). As green bioreactors, plants offers a variety of advantages such as nearly unlimited scalability, from small scale trials in growth chambers to large open-field mass production, and all at relatively inexpensive cost. Plants have become a promising alternative over the traditional expression systems to produce a variety of valuable biological molecules ranging from medicinal applications such as vaccines to materials like biodegradable plastics with industrial uses  (Twyman  et al.,  2005) .Plants can produce sufficiently high yields of proteins than bacterial or yeast fermentation systems and at 0.1% of the cost of mammalian cell cultures (Twyman...

  Types of Biomolecules and recent examples In recent years, several important products such as human biopharmaceuticals, recombinant antibodies, recombinant subunit vaccines, nutritional supplements, biodegradable plastics have been produced in plants with high success (Miao  et al ., 2008). The first pharmaceutically relevant protein made in plants was human growth hormone, which was expressed in transgenic tobacco in 1986 (Barta  etal ., 1986). The first antibody was also expressed in tobacco in 1989,which proved that plants could assemble complex functional glycoproteins with several subunits (Hiatt  et al ., 1989). Since then, other important vaccine candidates and therapeutic proteins have been produced in transgenic plants and are in different stages of clinical trials (Ma  et al ., 2003). Some important recombinant products produced in plant systems is given in table 2.  Plants can be engineered to act as bioreactors for vaccines and therapeutic pro...

  Plant made pharmaceuticals (PMPs) Therapeutic proteins are bioactive molecules that have potential applications in medicinal diagnostics and therapy. Several therapeutic products can be produced in plants which include diagnostic proteins (antibodies and enzymes), replacement proteins (Factor VIII for hemo philiacs, in sulin for diabetics), immune system stimulator/suppressants (interleukins, interferons, and colony stimulating factors), and adhesive proteins for surgical purposes or for growth factors (Daniell  et al.,  2001b; Goldstein and Thomas, 2004; Rajasekharan, 2006; Twyman  et al ., 2005). Antibodies or immunoglobulins (IgGs) are serum proteins that play a central role in the humoral immune response and the production of these antibodies in plants are referred as “Plantibodies” (De Jaeger  et al.,  2000; Goldstein and Thomas, 2004). The production of immunoglobulin fragments and their assembly in plants was reported in tobacco for the first time ...

  Plant made nutritional compounds (PMNs) Plants can provide most of the nutrients required in the human diet. However, major crops have been found to be deficient in one or the other nutrients. The advances in genetic modifications have made it possible to enhance the nutritional quality of the plants (Galili  et al ., 2002; Zimmermann and Hurrell, 2002). Several technical advances have been made from earlier attempts to simultaneously manipulating multiple steps in plant metabolic pathways and in constructing novel, multi-enzyme pathways in plant tissues (Kinney, 2006; Sandmann  et al ., 2006; Wu  et al ., 2005). In the last few years, a lot of progress has been made in the field of biofortification, specifically palnts with higher β-carotene, lycopene, vitamins, flavonoids, resveratrol, polyamines, nutraceuticals, amino acids, nutritional proteins, minerals, fatty acids, and carbohydrates are being produced.

  Plant made vaccines (PMVs) A vaccine is an antigenic preparation used to establish immunity against a disease and the main aim of the vaccination is to eradicate infectious diseases. In the beginning (Prakash, 1996; Artnzen, 1997)proposed that plants can be engineered for the production of vaccines. Several antigenic determinants belonging to various pathogens causing variety of diseases including bacterial and viral diarrhea, anthrax, rabies, cancer, SARS, measles, HIV, diphtheria, pertusis, tetanus, tuberculosis, respiratory syndrome, Alzheimer's disease, malaria, foot and mouth disease of cattle, gastroenteritis, hemorrhagic disease, bursal disease, goat plague, rinder pest virus, cytomegalovirus infections, parvoviral infections of dogs, avian influenza and bovine pneumonia have been produced in plants (Khandelwal  et al.,  2003; Sharma  et al.,  2004; Streatfield and Howard, 2003; Tiwari  et al.,  2009; Youm  et al.,  2008). In recent ...

  Plant made plastics (PMPs) Plastics difficult to dispose off and continually accumulating non-degradable wastes have become a significant source of environmental pollution (Shimao, 2001). Biodegradable plastics seem to be a viable alternative to synthetic plastics. The biodegradable materials canundergo decomposition into carbon dioxide, methane, water, inorganic compounds with the help of the enzymatic actions of microorganisms within a specified period of time (Anderson and Dawes, 1990). Polyhydroxyalkanoates (PHAs) are the biodegradable polymers which occur naturally in plants. Plants have been engineered to produce PHAs or PHBs in the various plant cell compartments (John and Keller, 1996; Matsumoto  et al.,  2009). For the economic feasibility of transgenic plants-derived biodegradable plastics, accumulation of at least 15% of the tissue dry weight is required (Scheller and Conrad, 2005). The expression level of biodegradable plastic-like compounds in plants, have ...

  Plant made industrial compounds (PMIs) Plant molecular farming is starting to become a viable new industry. This group includes hydrolases, encompassing glycosidases and proteases, milk proteins ß-casein, lactoferrin and lysozyme, protein polymers tissue replacement (Ma  et al.,  2003). Expression of thioredoxin in foods such as cereal grains would increase the digestibility of proteins and thereby reduce their allergenicity (Thomas  et al ., 2002). Human collagen can be produced in transgenic tobacco plants and that the protein is spontaneously processed and assembled into its typical triple-helical conformation (Ma  et al.,  2003). The production of chicken egg white avidin in transgenic corn using an avidin gene with codonoptimization was achieved (Hood  et al.,  1997). The endoplasmic reticulum of transgenic tobacco and vacuole of potato tubers expressed recombinant dragline silk protein up to 2% of TSP (Scheller  et al.,  2001). S...

  The plant endomembrane system A plant cell contains upto 10,000 different kinds of proteins. Each of these proteins must be localized to the precise intracellular membrane, organelle or directed to the cell surface for its proper functioning. The plant endomembrane system is a complex system of organelles specialized for the synthesis, transport, modification and secretion of proteins and other macromolecules. This system is composed of several functionally distinct membrane compartments: the endoplasmic reticulum (ER), the Golgi apparatus including the trans-Golgi network (TGN), secretory vesicles, the vacuole/lysosome andendosomes. Notably, the membranes of mitochondria and chloroplasts do not belong to the endomembrane system (Vitale and Galili, 2001; Dacks  et al ., 2009). However, the synthesis of the majority of proteins of a eukaryotic cell occurs in the cytosol, and from there proteins are migrated to reach their final destination. These proteins thus contain the inf...

  Subcellular Targeting of proteins in plants Organelle-specific protein targeting, protein sequestration in, or targeting to a specific cell compartments has also been readily recognized as a key factor determining the overall stability and yield of recombinant proteins in plants (Wandelt  et al.,  1992; Schouten  et al.,  1996; Gomord  et al.,  1997). Targeting signals can be used to intentionally retain recombinant proteins within distinct compartments of the cell to protect them from proteolytic degradation, preserve their integrity and to increase their accumulation levels (Seon  et al.,  2002). Several subcellular compartments have been considered as possible destinations for recombinant proteins in plant cells, endoplasmic reticulum, chloroplast and different subcompartments of the cell secretory pathway (Ma  et al.,  2003; Daniell, 2006; Goulet and Michaud, 2006). Recombinant products and their localization sites in the plan...

  Different plant species as platform for molecular farming The range of plant species amenable to transformation is growing at a phenomenal rate and it is unclear at present which species are optimal for molecular farming. Many factors need to be taken into consideration (Schillberg  et al ., 2003). The factors that are taken into consideration are the total biomass yield, the storage and distribution of the product. Various production platforms have been developed for molecular farming in plants which includes leafy crops (alfalfa, lettuce,  Arabidopsis , spinach, tobacco), cereals and legumes (barley, maize, pea,pigeon pea, rice, wheat), fruits and vegetables (banana, carrot, potato, tomato, carrots), oil yielding plants (false flax, flax, rape, safflower, soybean, white clover, white mustard) and sugar crops (sugar beet and sugarcane) (Twyman  et al ., 2003 and 2005).   ·       Tobacco Tobacco have well developed technology for gene ...

  Plant Transformation Two transformation approaches are commonly used to produce recombinant pharmaceuticals in plants (i) Transient expression and ii) Stable transformation of crop species. There are three major transient expression systems to deliver a gene to plant cells includes delivery of projectiles coated with ‘naked DNA’ by particle bombardment, infiltration of intact tissue with recombinant  Agrobacterium  (agro infiltration), or infection with modified viral vectors. Stable expressions of transgenes include insertion of genes in the nuclear genome of transgenic plants by two general methods Agrobacterium -mediated transformation and particle bombardment.   Transient expression in plants The transient production platform is perhaps the fastest and the most convenient production platform for plant molecular farming (Rybicki, 2010). The systems, which are mainly used for quick validation of expression constructs, are routinely used for the production of cons...