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Vector Incert Size

Vectors are DNA molecules that can replicate inside a host cell and integrate foreign DNA. Plasmids were the first vectors used to create recombinant DNA, allowing for the transfer of DNA between bacteria. Plasmids are extrachromosomal, circular DNA molecules commonly used as vectors due to their small size, high copy number, and ability to carry foreign DNA. Choice of vector depends on insert size, vector size, restriction sites, and cloning efficiency.

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31 views4 pages

Vector Incert Size

Vectors are DNA molecules that can replicate inside a host cell and integrate foreign DNA. Plasmids were the first vectors used to create recombinant DNA, allowing for the transfer of DNA between bacteria. Plasmids are extrachromosomal, circular DNA molecules commonly used as vectors due to their small size, high copy number, and ability to carry foreign DNA. Choice of vector depends on insert size, vector size, restriction sites, and cloning efficiency.

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maruthisreenivas
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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

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