VECTOR

DEFINITION : A DNA molecule that has the ability to replicate inside the host
to which the desired gene is integrated for cloning

Ex : Plasmids, Bacteriophages, Cosmids, BAC, YAC, Shuttle vectors, PBR

322 etc.

choice of Vector is determined basing on the following factors :

1) Incert size

2) Vector size

3) Restriction sites

4) Cloning efficiency

Vector Incert size

Plasmid < 10 kb

Bacteriophage 9 – 15 kb

Cosmid 33 – 50 kb

YACs 100 – 1000 kb

 PLASMIDS
In 1972 Paul berg along with Boyer and Cohen incorporated a DNA fragment
into a Plasmid creating the first Recombinant molecule. Later this was
transformed into a bacterium creating the first genetically engineered
organism.

Even now plasmids play a major role as vectors in DNA technology.

• Plasmids are extra chromosomal, double stranded, circular DNA

molecules present in bacteria

• Plasmids are either stringent or relaxed

• If the plasmid occurs free in cytoplasm then it is called a relaxed as it

can replicate independent of bacterial genome replication

• If the plasmid is integrated to the bacterial chromosome then it is called

as episome.
• Such plasmid can replicate only when bacterial genome replicates and
called as stringent.

• Relaxed plasmids are used as vectors for gene transfer in rDNA

technology

• Plasmids encode genes for specific sequence

• Types : 1) F-plasmids 2) R-plasmids 3) Col-plasmids 4) Metabolic

plasmids

• F-plasmids ( fertility factor or sex factor)

• Bacterial cells with F-plasmids are called F+ cells and acts as Donor
cells or +strain during bacterial conjugation. Ex. F plasmid of E.coli

• R-plasmids ( Resistance plasmids)

• These plasmids encode genes that confer bacterium resistance against

antibiotics. R plasmids are easily transferred from one strain to another
offering antibiotic resistance to neighbouring cells easily. Ex. RP4
plasmid of pseudomonas

• Col-plasmids : these plasmids carry genes that produce bacterial toxins

called colicins

• Ex. Col EI plasmid of E.coli

• Metabolic plasmids : carry genes involved in the production of metabolic

enzymes

• Ex. TP4 plasmid of pseudomonas

 Characteristics of Plasmids
1) smaller in size

2) high copy number

3) visual marker

4) promoters for in vitro transcription

Limitation of cloning in Plasmids

• Upper limit for clone DNA size is 10 kb

• Requires the preparation of “competent” host cells

• Insufficient for generating genomic libraries as overlapping regions

needed to place in proper sequence

• Preference for smaller clones to be transformed

• If it is an expression vector there are often limitations regarding

eukaryotic protein expression