BY: RAHUL ANDHARIA (MSIWM001)
Deals with insertion of desirable characteristics into plants through genetic modifications for the purpose of creating beneficial plants. Plants which are modified genetically are termed as Transgenic plants. Transgenic plants usually are created by modifying their DNA to serve different purposes.
The principle foundation of plant biotechnology was through the theories of cellular Totipotency( totipotency is the ability of single cell to divide and differentiate) and Genetic transformation. This theories led to the development of modifications in plants. Genetic transformation theory was proposed by Griffith, while cell theory was given by Schleiden and Theodore Schwann.
The plant breeding process has a rich history and developments in plants began as early as 12000 years ago. It is known that, the bread making wheat was started in Egypt during 4000-5000bc where it was grown in Egypt and cultivated in China. Selective breeding techniques were known to be used first by Babylonians for date palm cultivation.(875bc). The term biotechnology in the year 1919 was first given by Karoly Ereky. Demonstration of plant tissue culture technique for the first time was given by Haberlandt.
Methods in Plant biotechnology:
Tissue and cell culture:
This technique helps to maintain cells and tissues for a longer duration of time in an artificial medium. Cells are isolated and grown in a medium which is called as tissue culture medium that contains all the necessary nutrients required for the plant growth and it’s proliferation.
- Selection of a suitable explant and than sterilizing it using disinfectant.
- Preparation of an appropriate culture medium which suits the explant selected.
- The prepared medium is than sterilized by using autoclave.
- Inoculation of the sterilized explant and is than transferred to the culture medium in aseptic conditions.
- Incubation of cultures in a culture room with proper temperature, moisture and humidity conditions.
- Next step is, sub culturing where the cultures are transferred to a fresh nutrient medium to obtain plantlets.
- After acclimatising plantlets to environmental conditions, they are transferred to green house or can be grown in pots.
By using this technique it is possible to cultivate any plant species by using a variable explant, for example: pedicle, stem segment, leaf segment, petiole, anther, etc. Plant tissue culture can be carried out in both solid or liquid media depending on the plant specie and requirement of conditions.
Recombinant DNA technology:
Technology in which DNA from one genome is inserted to the other. This approach is best suited for recombination between unrelated species. With this technique, foreign gene is introduced to the plant genome artificially and the resultant plant is genetically engineered or modified.
Three main components involved with genetic engineering:
- Isolation of foreign gene from a suitable source.
- Suitable vector carrying the gene.
- Various means to introduce the vector into the host genome.
Genetic Engineering has following steps:
Isolation of gene of interest to be cloned from desired organism.
Transfer of the isolated gene to create recombinant DNA molecule. By cutting the DNA molecules at specific sites by using Restriction Enzymes, recombinant DNA molecules can be created.
Restriction enzymes cut DNA at specific locations. Restriction enzymes are designated as I, II, III, and IV. This enzymes varies in their structure, site of cleavage and some cofactors (substance which is required for enzyme activity).
By the action of RE, DNA is left with sticky ends (short portion of unpaired bases). Same restriction enzymes is used to cut the plasmid, so that the plasmid also has the similar sticky ends and can base pair.
The plasmid and the isolated gene are joined together by an enzyme called DNA ligase.
The two pieces of DNA with same sticky ends (as cut by same RE) are being linked together by DNA ligase and forms a single, unbroken molecule.
Than the genetically engineered plasmid is inserted into bacterial cell (Transformation). The gene is transferred by means of Vectors. Selection of vectors is specific and will depend on the type of gene to be inserted.
When the bacteria reproduce, the plasmid will get copied and this recombinant plasmid spreads as bacteria multiply and expresses the gene and makes a human protein.
By using recombinant DNA technology, transgenic plants can be prepared.
Plants with their genome modified by either addition of a foreign gene or removal of damaged gene are called as Transgenic plants. Transgenic plants are created based on the need. For example plants can be modified to resist pests, pathogens, insects and environmental conditions.
Example: Bt cotton is a genetically modified crop. It is modified to combat bollworms.
Production of transgenic plants using Ti plasmids:
Agrobacterium tumifaecins, is a soil bacterium that causes crown gall disease by incorporating the T-DNA region of Ti(tumour inducing plasmid) into the host cells. Thus, Ti plasmids can be used to prepare genetically engineered plants by modifying the T-DNA region of Ti plasmid. Steps are as follows:
- Ti plasmid is genetically engineered at T-DNA region by inserting an antibiotic resistant gene.(Kan R- kanamycin) as well as a foreign gene of interest.
- The plant cells in culture containing this cointegrated Ti plasmid transfers the foreign DNA into host cell.
- Thus, when foreign DNA is integrated, it disrupts the tumour formation, and only those plant cells can grow which are resistant to Kan-R.
- Plants are than regenerated from the culture through calluses.
- Foreign gene is usually expressed by the adult transgenic plants.
- This method is also called as variation breeding.
- In this method the seeds are exposed to radiation or chemicals to generate mutants.
- This technique generally helps to improve disease resistance to plants.
- It helps to improve specific characteristics of high yielding varieties. For example- Jiahezazhan Rice.
- Mutation can be induced by both physical and chemical agents.
- This type of breeding is more suitable to improve one or two specific traits. (Selective traits)
Crossing between 2 different plants to produce hybrids is called as hybrid breeding. Hybrid breeding is generally used to create hybrids that are completely different product as they are made from different parent lines.
Advantages of hybrid breeding:
- Fast growing plants, so it is advantageous for farmers as they can reap more crop and can earn larger profit.
- More disease resistant than parent plant.
- Can withstand abiotic stresses.
Disadvantages of hybrid breeding:
- Heterosis effect( progeny exhibits greater fertility, biomass, growth than parents) lasts only for one generation.
Application and Potential of Plant biotechnology:
- Large scale plant species can be raised by this method.
- Meristem is used in this method and is cultured in basal medium containing hormones, nutrients, carbohydrates and nitrogen sources.
- Technique is employed to eliminate pathogens and viruses.
- Examples of plants: Sugarcane and potato are micro-propagated commercially to prevent virus and pathogen free plants for better yield and profit.
Herbicide resistant plants:
- Weeds often ground the crop plants and reduces the yield. To control weeds, farmers uses herbicides to destroy weeds.
- But generally, it has been observed that herbicides used can cause side effects to plants. For this reason genes for some enzymes have been genetically engineered to provide resistance to various herbicides. Table below is attached showing the gene strategy used and the plants benefitted through it:
Resistant to abiotic stresses:
- Conditions like drought, salinity(high salt concentration), flooding, heat and freezing leads to poor harvest of the crop.
- Protective proteins or enzymes from other plants or organisms are genetically engineered to fight against such adverse conditions.
- saturation levels of membrane fatty acids are usually altered. Other than this Osmolytes, osmoprotectants and rate of reactive oxygen intermediate is also changed.
- Abiotic stress tolerant genes, when introduced into plants can provide tolerance to abiotic stresses.
- Example- Enzyme choline dehydrogenase from E.coli when injected into potato and tobacco plants, produces glycine betaine, which resistance to salts and freezing.
Insect pests and disease resistance:
- Insects destroys plants and leads to economic loss. Traditional insecticides can kill the pests but simultaneously it also causes damage to useful insects and leads to soil damage.
- Many strategies have been adopted to engineer disease resistant gene in plants. One common example is Bt cotton, where resistant gene is introduced to kill cotton bollworms.
- Genes conferred with protease inhibitors, alpha amylases are introduced which interacts with insect metabolism and confers resistance by destroying it.
Apart from these, plant biotechnology is also used to improve nutritional quality of food and to enhance its nutritional value. Nutritional value of plants can be increased by altering the amino acid composition of plants proteins and by introducing transgene with desired traits.