Types of cloning Vectors for gene transfer

plant gene transfer

1. Vectors
•Cloning vectors are DNA molecules that
are used to "transport“ DNA sequences
between biological hosts and the test tube.
•Most vectors are genetically engineered.
•A vector is used to amplify a single
molecule of DNA into many copies.

2. Plasmids: VarietiesPlasmids: Varieties
Relaxed plasmids or high copy number plasmids
Stringent plasmids or low copy number plasmids
oNarrow-host range plasmids
oWide host range plasmids
Naturally occurring plasmids
Synthetic plasmids.
Based on copy numbers
Based on host range
Based on occurence

3. Col plasmids: Col plasmids contain genes that make bacteriocins (aka
colicins), which are proteins that kill other bacteria and thus defend the host
bacterium.
Resistance (R) plasmids: Resistance or R plasmids transfer through
conjugation and contain genes that help a bacterial cell defend against
environmental factors such as poisons or antibiotics.
Fertility (F) plasmids: Fertility plasmids, also known as F-plasmids fall under
the broad category of conjugative plasmids and contain transfer genes that
allow genes to be transferred from one bacteria to another through conjugation.
Virulence (Vir) plasmids: A virulence plasmid turns a host bacterium into a
pathogen, which is an agent of disease.
Degradative plasmids: These help the host bacterium to digest compounds
that are not commonly found in nature, such as camphor, xylene, toluene, and
salicylic acid
Based on specific function/role

4. Bacteriophage
Bacteriophages are a group of viruses that used
bacterial cells as host and reproduce by infecting
bacterial cells.
Like all other viruses bacteriophages are also
consisting of a protein coat that provides protection to
the genome.
Most of the phages are DNA viruses that code for
several genes responsible for virus replication.

5. Types of phage vectorsTypes of phage vectors
20-25kb insert capacity

6. CosmidsCosmids
Plasmid derived vector - containing cos sites from phage are known as
cosmids.
Reported by Collins and Hohn, 1978.
Cosmids can be used to clone large fragments of DNA by exploiting the
phage in vitro packaging system.

7. Phasmids (Phagemids)
•A
 
phagemid 
or 
phasmid 
is a DNA-based cloning vector, which has
both
 bacteriophage and plasmid properties. 
•They carry an
 origin of replication derived from bacteriophage (f1) in
addition to origin of plasmid replication.
•Phagemid vectors differ by having the ability to be packaged into
the
 capsid of a bacteriophage, due to their having a genetic
sequence that signals for packaging.
•Phagemids are used in a molecular biology technique called
"Phage Display“ (to study protein-protein interactions
 and other
ligand/receptor combinations.)

8. Shuttle vectors
•A
 shuttle vector is a vector (usually a plasmid)
constructed so that it can propagate in two different
host species.
•DNA inserted into a shuttle vector can be tested or
manipulated in two different cell types.
•Main advantage of these vectors is they can be
manipulated in
 
E. coli, then used in a system which is
more difficult or slower to use (e.g. yeast).
•Shuttle vectors can propagate in
 eukaryotes and prokaryotes 
(e.g. both
 
Saccharomyces cerevisiae 
and 
Escherichia coli) or
in different species of bacteria (both
 
E. coli 
and 
Rhodococcus
 erythropolis
).
There are also
 adenovirus shuttle vectors, which can propagate in 
E. coli 
and
mammals.

9. Yeast Artificial Chromosome (YAC) is a vector used to clone
DNA fragments larger than 100 kb and up to 3000 kb.
First described in 1983 by Murray and Szostak, a YAC is an
artificially constructed chromosome that contains a
centromere, telomeres and an autonomous replicating
sequence (ARS) element, which are required for replication
and preservation of YAC in yeast cells.
YACs are useful for the physical mapping of complex
genomes and for the cloning of large genes.

10. PAC – P1 derived Artificial Chromosome
PAC, is a DNA construct derived from the DNA of
P1 bacteriophages and BAC.
Efficient cloning vector to clone DNA fragments
(100- to 300-kb insert size; average,150 kb) in
E coli cells.
P1 was developed as a cloning vector by
 
Nat Sternberg
 
and colleagues in the 1990s

11. Applications of PAC
building genome libraries for human, mouse, etc, helps
with projects such as Human Genome Project
libraries served as the template for gene sequencing
genome analysis on specific functions of different genes for
more complex organisms (plants, animals etc.)
facilitate gene expression