Recombinant DNA Technology (a new approach in Biotechnology) Dr. O. S. Deshmukh Assistant Professor in Botany Mahatma Fule Arts, Commerce and Sitaramji Chaudhari Science Mahavidyalaya, Warud, Dist. Amravati. 2018

Index Brief History Principle of Recombinant DNA Technology. Molecular Tools of r-DNA Technology What is Restriction EndoNuclease? Nomenclature Recognition Sequences Cleavage Pattern What is DNA Ligase?

Index What are Host Cells? What is Vector? What is Plasmid? What is Transformation? What are the applications of r-DNA Technology? Conclusion. REFRENCES......

The Recombinant DNA speaks “I am the hybrid DNA molecule, Created by cutting and sealing, When introduced into the host cell, I multiply and code for the desired proteins.....”

Brief History The present day DNA technology has its roots in the experiment performed by Boyer and Cohen in 1973. 1. BBC Biotechnology – Before Boyer and Cohen. 2. ABC Biotechnology – After Boyer and Cohen.

Principle of Recombinant DNA Technology Generation of DNA fragments and selection of the desired piece of DNA.

Principle of Recombinant DNA Technology Insertion of the selected DNA into a Cloning vector

Principle of Recombinant DNA Technology Introduction of Recombinant vector into host cells

Principle of Recombinant DNA Technology Multiplication and selection of clone and Expression of the gene to produce the desired product

Molecular Tools of r-DNA Technology - Restriction EndoNuclease – DNA cutting enzyme DNA Ligase – DNA Sealing enzyme

What is Restriction EndoNuclease? The bacterial enzymes that can cut/ Split DNA at specific sites. Restriction EndoNuclease was first discovered in E. coli, restricting the replication of Bacteriophage, by cutting the viral DNA. They are known as restriction enzymes or restriction EndoNuclease or Genetic Scissor.

Nomenclature - The first letter of the enzyme indicate the genus name, followed by the first two letter of the species, then comes the strain of the organism and finally roman numerical indicating the order of discovery. E.g. EcoRI is from Escherichia (E) coli (co) Strain RY-13 (R) and first EndoNuclease (I) Hind III Haemophilus (H) influenzae (in) Strain Rd (d) and third endonuclease (III)

Recognition Sequences - It is the site where the DNA is cut by restriction endonuclease. Restriction endonuclease can specifically recognize DNA with particular sequence of four to eight nucleotides and cleave.

Cleavage Pattern -

What is DNA Ligase? DNA Ligase actively participates in cellular DNA repair process. DNA Ligase joins (Seals) the DNA fragment by forming a phosphor di ester bond

What are Host Cells? Factories of cloning The living system or cells in which the carrier of recombinant DNA molecule or vector can be propagated. Prokaryote (Bacteria) and Eukaryote (Fungi, animal and Plants)

What is Vector? Cloning Vehicle DNA molecule which can carry a foreign DNA fragment They are self replicating E.g. Plasmids, Bacteriophage, Cosmids and artificial chromosome vectors.

What is Plasmid? Extra chromosomal double stranded circular self replicating DNA molecules. All the bacteria have plasmids. The size of plasmid vary form 1 – 500 kbp.

What is Transformation?

What are the applications of r-DNA Technology? 1. Genetic Engineering. 2. Blotting Technique. 3. DNA sequencing. 4. DNA Chips. (Micro arrays) 5. Polymerase Chain Reaction (PCR) 6. Gene libraries - a computerized listing of known DNA sequence.

Gene Library

What are the applications of r-DNA Technology? 7. Gene Bank. 8. Insulin and Diabetes control. 9. Recombinant vaccines. 10.DNA Vaccines. 11.Transgenic Plants and Animals. 12.Gene Therapy

Conclusion – 1. r-DNA technology is primarily with the manipulation of genetic material (DNA) to achieve the desired goal in a predetermined way. 2. Provide strategies of gene cloning.

References – Biochemistry – U. Satynarayana and U. Chakrapani. Plant Biotechnology – K. G. Ramawat Biotechnology – 3 – M. K. Sateesh The Cell – Cooper www.sinauer.com www. biology.arizona.edu.co.in

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