Recombinant DNA technology involves combining DNA from different sources, such as combining an animal gene with bacterial DNA, to create recombinant DNA with new functions. Key discoveries that enabled recombinant DNA technology include determining the DNA structure, and isolating enzymes like DNA ligase and restriction enzymes. The goals of recombinant DNA technology include isolating and characterizing genes, altering genes, and returning altered genes to living cells. The process involves isolating DNA, cutting DNA with restriction enzymes, joining DNA together, amplifying the recombinant DNA in bacterial hosts, and using vectors like plasmids, cosmids, and phages to replicate and select for recombinant DNA. Applications include producing medicines like insulin through gene therapy and agriculture through genetically modified crops.

1. 1
Presented by Rufia Abbas
Recombinant DNA technology

2. Recombinant DNA technology
DNA molecules that are extracted from different
sources and chemically joined together; for example
DNA comprising an animal gene may be recombined
with DNA from a bacterium.
This method can be used to combine DNA from
different species or to create genes with new
functions. The resulting copies are called
recombinant DNA.
2

3. Discovery of recombinant DNA
technology
Discovery of DNA structure Watson & Crick in 1953
Isolation of DNA ligase in 1967
Isolation of REase in 1970
Paul Berg generated rDNA technology in 1972
Cohen & Boyer in 1973 produced first plasmid vector
capable of being replicated within a bacterial host
3

4. Goals of recombinant DNA technology
• To isolate and characterize a gene
• To make desired alterations in one or more isolated genes
• To return altered genes to living cells
• Artificially synthesize new gene
• Alternating the genome of an organism
• Understanding the hereditary diseases and their cure
• Improving human genome
4

5. Procedure of making rDNA
Isolating of DNA
Cutting of DNA
Joining of DNA
Amplifying of DNA
5

6. Isolating of DNA 6

7. Cutting of DNA
• DNA can be cut into large fragments by mechanical
shearing.
• Restriction enzymes are the scissors of molecular
genetics.
The Agarose Gel electrophoresis technology
display the restriction enzyme digestion
progress.
7

8. Joining DNA 8

9. Amplifying the recombinant DNA
• Transforming the recombinant DNA into a bacterial host strain.
• The cells are treated with CaCl2
• DNA is added
• Cells are heat shocked at 42 C
• DNA goes into cell by a somewhat unknown mechanism.
• Once in a cell, the recombinant DNA will be replicated.
• When the cell divides, the replicated recombinant molecules go to both
daughter cells which themselves will divide later. Thus, the DNA is
amplified
9

10. Vectors used in rDNA technology
• A vector is an area of DNA that can join another DNA
part without losing the limit for self-replication
• Should be capable of replicating in host cell
• Should have convenient RE sites for inserting DNA of
interest
• Should have a selectable marker to indicate which
host cells received recombinant DNA molecule
• Should be small and easy to isolate
10

11. Vectors used in rDNA technology
BACS
YACS
plasmid
vectors
cosmid
Lamda
phage
express
ion
11

12. Applications of rDNA technology
• Agriculture: growing crops of your choice (GM food),
pesticide resistant crops, fruits with attractive colors, all
being grown in artificial conditions
• Pharmacology: artificial insulin production, drug delivery to
target sites
• Medicine: gene therapy, antiviral therapy, vaccination,
synthesizing clotting factors
• Other uses:fluorescent fishes, glowing plants etc
12

13. References
• Campbell, Neil A. & Reece, Jane B.. (2002). Biology (6th ed.). San
Francisco: Addison Wesley.
• Peter Walter; Alberts, Bruce; Johnson, Alexander S.; Lewis, Julian; Raff,
Martin C.; Roberts, Keith (2008).
• Berg, Jeremy Mark; Tymoczko, John L.; Stryer, Lubert (2010).
• Russell, David W.; Sambrook, Joseph (2001). Cold Spring Harbor, N.Y:
Cold Spring Harbor Laboratory.
• Hannig, G.; Makrides, S. (1998). "Strategies for optimizing heterologous
protein expression in Escherichia coli".
• Mahmoudi Gomari, Mohammad; Saraygord-Afshari, Neda;
Farsimadan, Marziye; Rostami, Neda; Aghamiri, Shahin; Farajollahia,
Mohammad M. (1 December 2020).
• Brondyk, W. H. (2009). "Guide to Protein Purification, 2nd Edition.
13

14. 14