Introduction, definition, types , properties, application and example pBR322 with a neat diagram

1. CLONING VECTORS
AASIA SULTANA M.

2. INDEX
 INTRODUCTION
 PROPERTIES OF CLONING VECTORS
 TYPES OF CLONING VECTORS
pBR322 PLASMID VECTOR

3. INTRODUCTION
• A cloning vector is a
small piece of DNA
that can be stably
maintained in an
organism into which
a foreign DNA
fragment can be
inserted for cloning
purposes.
• A cloning vector may
be DNA taken from a
virus/cell of a higher
organism.

4. WHY CLONING VECTORS ARE USED?
Used as a vehicle to artificially carry foreign genetic material into
another cell where it can be replicated and expressed.
Amplify a single molecule of DNA  Many copies [clones].
Used to transport cloned sequence of desired gene.

5. PROPERTIES
1. Capable of autonomous replication within a host cell.
2. Contains at least one restriction endonuclease site.
3. Carries at least one gene that can be used to select the cloned vector such as an
antibiotic resistance gene (selectable markers).

6. CLONING VECTORS
• Plasmids
• Phages
• Cosmids
• Bacterial Artificial Chromosomes
(BAC).
• Yeast Artificial Chromosomes (YAC).
• Human Artificial Chromosomes (HAC).
• Shuttle and Expression Vectors.

7. TYPES OF CLONING VECTORS
PLASMIDS
• Plasmids are extra chromosomal circular double stranded DNA replicating elements
in bacterial cells.
• Plasmids show the size ranging from 5.0 kb to 400 kb.
BACTERIOPHAGE
• The viruses that infect bacteria are called bacteriophage. These are intracellular obligate
parasites that multiply inside bacterial cell by making use of some or all of the host
• Bacteriophages have a very high significant mechanism for delivering its genome into
bacterial cell. Hence it can be used as a cloning vector to deliver larger DNA segments.

8. BACTERIAL ARTIFICIAL CHROMOSOMES
• Bacterial artificial chromosomes (BACs) are simple plasmids designed to clone very large DNA fragments ranging from 75 to 300 kb in size.
• BACs basically have marker like sights such as antibiotic resistance genes and a very stable origin of replication.
YEAST ARTIFICIAL CHROMOSOMES
• A very large DNA fragments whose sizes ranging from 100 kb to 3000 kb can be cloned using YACs.
• Used for physical mapping of complex genomes.
HUMAN ARTIFICIAL CHROMOSOMES
• HACs are microchromosomes that can act as a new chromosome in a population of human cells.
• HACs range in size from 6 to 10 Mb that carry new genes introduced by human researchers.
SHUTTLE & EXPRESSION VECTORS
• Used for synthesis and expression of proteins coded by inserted DNA.
COSMIDS
• These are circular extra chromosomal DNA molecule combining the features of plasmids and phages.

9. Features:
1. Ori – Origin of replication:
Replication becomes fast and number of copies can be
decided.
2. Cloning sites:
Restriction enzymes acts on this sites. There must be one site
for each RE.
3. Selectable markers:
Genes which helps to differentiate transformant and non
transformant.
pBR 322 PLASMID VECTOR

10. • pBR 322: p-Plasmid, B-Boliver, R-Rodriguez and 322-
Plasmid’s number.
• Plasmids have double stranded circular DNA and
extra chromosomal DNA.
• It is an artificially synthesized vector from E.coli.
• Two special genes ampR-Ampicillin Resistance
and tetR-Tetracycline Resistance.
• Puv I, Pst I, Puv II, Sal I, BamH I, Hind III, Cla I and
EcoR I are restriction endo nucleases.