Plant transformaation technology, all techniques

Introduction

Why transform plants

Plant transformation technique

Agrobacterium mediated transformation

Biolistics method

Microinjection method
Conclusion
References

1. Plant transformation technology By KAUSHAL KUMAR SAHU Assistant Professor (Ad Hoc) Department of Biotechnology Govt. Digvijay Autonomous P. G. College Raj-Nandgaon ( C. G. )

2. synopsis Introduction Why transform plants Plant transformation technique Agrobacterium mediated transformation Biolistics method Microinjection method Conclusion References

3. Plant transformation is the introduction of a foreign piece of DNA, confering a specific trait, into host plant tissue. The foreign gene (termed the " transgene ") is incorporated into the host plant genome and stably inherited through future generations. Plant transformation can be carried out in a number of different ways depending on the species of plant . Plant transformation was developed as an alternative to conventional breeding methods which are more laborious than this fairly simple (now routine) laboratory procedure. Introduction

4. Why transform plants ??? Plants are transormed in order to create transgenic plants that contain genes conferring traits that are desirable to the plant breeder. This includes resistance to specific plant diseases and pests, drought and saline tolerance. It also includes novel uses of plants, such as plantibody production and the production of edible vaccines in plant tissue. Conventional breeding methods were seen to be slow and laborious, plant transformation techniques were developed to overcome these problems. The ability to introduce novel genes into plants, often across the species barrier, provides many commercially viable applications to the plant breeder.

5. Specific genes have been transferred into the plant to improve their various features, eg . Herbicide resistance. Several gene transfers have been aimed at improving the produce quality, eg . Protein or lipid quality. Transgenic plants may be developed to produce novel biochemicals like interferon, insulin, immunoglobulin, etc. Transgenic plants have been developed with a view to produce recombinant vaccines. A pplication of gene transfer

6. Plant Transformation Methods Physical Chemical Biological Microinjection Biolistics - gene gun Electroporation Microinjection Silica/carbon fibers Lazer mediated PEG DEAE- dextran Calcium phosphate A. Tumefaciens Rhizogenes Virus-mediated

7. Agrobacterium mediated transformation Agrobacterium tumefaciens is a widespread naturally occurring soil bacterium that causes crown gall, and has the ability to introduce new genetic material into the plant cell . During transformation, several components of the Ti plasmid enable effective transfer of the genes of interest into the plant cells. These include: T-DNA border sequences, which demarcate the DNA segment (T-DNA) to be transferred into the plant genome vir genes (virulence genes), which are required for transferring the T-DNA region to the plant but are not themselves transferred, and modified T-DNA region where the genes that cause crown gall formation are removed and replaced with the genes of interest.

8. Protocol The Agrobacterium -mediated transformation process involves a number of steps: isolation of the genes from the source organism; (b) development of a functional transgenic construct including the gene; promoters to drive expression and marker genes to facilitate tracking of the introduced genes in the host plant; (c) insertion of the transgene into the Ti-plasmid; (d) introduction of the T-DNA-containing-plasmid into Agrobacterium ; (e) mixture of the transformed Agrobacterium with plant cells to allow transfer of T-DNA into plant chromosome.

9. (f) regeneration of the transformed cells into genetically modified (GM) plants; and (g) testing for trait performance or transgene expression at lab, greenhouse and field level. Figure : Agrobacterium -mediated Plant Transformation Process

10. Biolistic method A gene gun or a biolistic particle delivery system , originally designed for plant transformation , is a device for injecting cells with genetic information. The payload is an elemental particle of a heavy metal coated with plasmid DNA . This technique is often simply referred to as bioballistics or biolistics . This device is able to transform almost any type of cell, including plants, and is not limited to genetic material of the nucleus: it can also transform organelles, including plastids .

11. Application Gene guns are so far mostly applied for plant cells. However, there is much potential use in animals and humans as well. Plants The use of the gene gun may be contrasted with the use of Agrobacterium tumefaciens and its Ti plasmid to insert genetic information into plant cells. The target of a gene gun is often a callus of undifferentiated plant cells growing on gel medium in a petri dish . After the gold particles have impacted the dish, the gel and callus are largely disrupted.

12. Humans and animals Gene guns have also been used to deliver DNA vaccines . The gene gun technique is also popularly used in an edible vaccine production technique, where the nano -gold particles coated with plant genetic material under the high vacuum pressurized chamber are transformed into suitable plant tissues.

13. Microinjection Microinjection  refers to the process of using a glass  micropipette  to insert substances at a  microscopic  or borderline  macroscopic  level into a single living  cell . It is a simple mechanical process in which a needle roughly 0.5 to 5 micrometers in diameter penetrates the  cell membrane . Microinjection is normally performed under a specialized  optical microscope  setup called a  micromanipulator .

14. The process is frequently used as a  vector  in  genetic engineering  and  transgenics  to insert  genetic material  into a single cell. Microinjection can also be used in the  cloning  of organisms, and in the study of cell biology and viruses.

15. Conclusion Plants are the first step in every food chain, therefore, humans and animals are dependent upon plants for their survival. Increases in plant production are needed to meet growing demands for food. Use of plant transformation techniques in plant biotechnology, including gene transfers into crop plants, offers significant advantages over traditional plant breeding in developing improved crops. Plant biotechnology is being used to produce plants capable of hyper accumulating toxic metals from soil and water, thus contributing to a cleaner environment.

16. References Plant biotechnology---- H.S. Chawla Textbook of biotechnology--- B.D. Singh www.wikipedia.com www.ncbi.com

17. Thankyou