1. CLONING VECTORS
Shumaila Azam

2. CLONING VECTORS
The DNA into which a foreign piece of DNA is cloned is called a ‘VECTOR’
Vectors are those DNA molecules that carry a foreign DNA fragment when inserted into it.
Vectors are also known as vehicle DNAs.
Based on the nature and sources the vectors are grouped into bacterial bacteriophages, plasmids, cosmids and phasmids.

3. TYPES OF VECTORS
If a vector is used for reproducing the DNA fragment, it is called a "cloning vector".  
If a vector is used for expressing certain gene in the DNA fragment, it is called an "expression vector".

4. CLASSES OF VECTORS Plasmids: Accept up to ~10 kb ‘foreign’ DNA
Phage : kb fragments (its own genome is only 50 kb!) Commonly used in making genomic libraries. (very high efficiency of transfection)
Cosmids: kb – similar to plasmids (high efficiency for transformations). Cosmids are hybrids of  phages and plasmids.
YACs (Yeast Artificial Chromosomes): kb! (essential for cloning very large fragments)

5. PLASMIDS

6. PLASMIDS The size of plasmids ranges from a few kb to near 100 kb.
Plasmids are self-replicating, double stranded, circular DNA molecules that are maintained in bacteria as independent extra chromosomal entities.
Virtually all bacterial genera have plasmids.
The size of plasmids ranges from a few kb to near 100 kb.

7. STRUCTURE OF PLASMID
 It contains a polylinker which can recognize several different restriction enzymes,
An antibiotic-resistance gene for selective amplification
A replication origin (ORI) for proliferation in the host cell.  

8. Structure of a typical plasmid

9. CLASSES OF PLASMIDS
Plasmids have basic attributes to make them potential vectors for carrying cloned DNA.
Some plasmids carry information for their own transfer from one cell to another called F plasmids.
Some plasmids encode resistance to antibiotics and are called as R plasmids.

10. CLASSES OF PLASMIDS
Some plasmids carry specific sets of genes for the utilization of unusual metabolites and are called as degradative plasmids.
Some plasmids have no apparent functional encoding genes and are called as cryptic plasmids.
Plasmids can range in size from less than 1 to more than 500kb.
Each plasmid has a sequence that functions as an origin of DNA replication, with out this site, it cannot replicate in a host cell.

11. CLASSES OF PLASMIDS
Some plasmids are represented by 10 to 100 copies per host ,these are called high copy number plasmids.
Some plasmids maintain 1 to 4 copies per cell and are called low copy number plasmids.
Seldom does the population of plasmids in a bacterium make up more than approx 0.1 to 5.0 percent of the total DNA.
When two or more types of plasmids cannot coexist in the same host cell, they are said to belong to a single incompatibility group.

12. CLASSES OF PLASMIDS
But plasmids from different incompatibility groups can be maintained together in the same cell. This coexistence is independent of the copy numbers of the individual plasmids
Some plasmids, because of their specificity of their origin of replication , can replicate in only one species of the host cell and are called narrow range plasmids.
Other plasmids have less specific origins of replication and can replicate in a number of bacterial species and are called broad range plasmids.

13. IMPORTANT FEATURES OF PLASMIDS
A small size, which is necessary because the efficiency of transfer of exogenous (foreign) DNA into E.coli decreases significantly with plasmids that are more than 15kb long.
Unique (single) restriction endonuclease recognition sites into which the insert DNA can be cloned
One or more selectable genetic markers for identifying recipient cells that carry the cloning vector --- insert DNA construct.

14. IMPORTANT FEATURES OF PLASMIDS
Consequently the plasmid cloning vectors have to be generally engineered.
However naturally occurring plasmids often lack several important features that are required for high quality cloning vector.

15. THANKYOU