A glance at Molecular Cloning

1. Molecular (Gene) Cloning
Mohammed Esawie, M.D., M.S.(c)

2. Background
• DNA amplification techniques produce
thousands or million copies of a particular
gene
• Either: In-vitro  PCR
In-vivo  Molecular Cloning
• In short: seek and find the gene  amplify it

4. Molecular (Gene) Cloning
• Definition??!
• Important components:
1- Target DNA
2- Restriction Endonucleases
3- Vectors
4- Marker gene
5- Recombinant DNA

5. On one side:
Target DNA & Restriction Endonucleases
• Secreted by bacteria  damage invading viral DNA
 Restricting its invasion
• Act on Restriction sites = palindromic Sequences
(4,6,8 bases read from 5’3’ in one DNA strand the
same as from 5’3’ in the antiparallel strand)
• EX:

6. On the other side:
Vector
• A molecule of DNA
• Contains:
Restriction site  will join to the gene to be
cloned
+ Marker gene 
to trace the vector after insertion
( to know which host cell accepts the vector) e.g
antibiotic resistance gene

7. On the other side:
Vector
• Types:
1- Prokaryotic Plasmids: extra-chromosomal
DNA (small, circular DNA.) e.g. E-coli plasmids
2- Viruses: naturally occurring viruses that infect
bacteria (Bacteriophage)
3- Cosmids: artificial recombinant plasmid can
insert larger DNA fragment up to 50 kb

8. Recombinant DNA
• Target DNA + Vector DNA
• Will be transferred from test tubes into host
cell that can provide the enzymatic machinery
for DNA replication (All DNA fragments
replication)

10. Molecular (Gene) Cloning Uses
• Gene study
• Synthesis of human protein drugs
• Vaccine preparation e.g. Hepatitis B Vaccine

11. Image Credits
• Amatakoon et al, DNA cloning and Restriction
Digestion. Pharmacognosy, 2007.
• Restriction enzymes & DNA ligase,
www.Khanacademy.org

12. Thank You