Plasmids are double-stranded DNA molecules that exist independently of the chromosome in organisms. They can replicate on their own and provide benefits to the host such as antibiotic resistance. Plasmids come in different types based on their ability to transfer horizontally or their function. Common plasmid types include F-plasmids, which allow for bacterial conjugation, and R-plasmids, which contain antibiotic resistance genes. Plasmids are also useful as vectors in biotechnology as they contain origins of replication, antibiotic resistance genes, and multiple cloning sites that allow for insertion and expression of foreign DNA.

1. Plasmids
Asheesh Kumar Mishra
DSMNRU Lucknow

2. Introduction
The word 'plasmid' was first coined by Joshua Lederberg in 1952.
Plasmids are double-stranded DNA molecules that can exist independently of the chromosome.
Extrachromosomal DNA molecules.
The genome of an organism includes all DNA, including plasmids.
Both circular and linear plasmids are documented.
Approx.: 30 genes.
Plasmids use the cell’s DNA-synthesizing machinery to replicate.
Replication not mentioned in any stage of cell cycle.
In some eukaryotic microorganisms, such as Saccharomyces cerevisiae.
Plasmids provide one or more functional benefits to the host such as resistance to antibiotics, degradative functions, and/or virulence.
Regulatory elements are also present at the ori site. For example- Rep proteins.

3. Structure

4. Vector Element Description
Origin of Replication (ORI) DNA sequence which allows initiation of replication within a plasmid by recruiting replication machinery proteins
Antibiotic Resistance Gene Allows for selection of plasmid-containing bacteria.
Multiple Cloning Site
(MCS)
Short segment of DNA which contains several restriction sites allowing for the easy insertion of DNA. In expression plasmids,
the MCS is often downstream from a promoter.
Insert Gene, promoter or other DNA fragment cloned into the MCS for further study.
Promoter Region
Drives transcription of the target gene. Vital component for expression vectors: determines which cell types the gene is
expressed in and amount of recombinant protein obtained.
Selectable Marker
The antibiotic resistance gene allows for selection in bacteria. However, many plasmids also have selectable markers for use
in other cell types.
Primer Binding Site
A short single-stranded DNA sequence used as an initiation point for PCR amplification or sequencing. Primers can be
exploited for sequence verification of plasmids.
Cont…

5. Classification
On the basis of their transfer ability
Conjugative Plasmid:
◦ Capable of horizontal transmission.
◦ Consist “tra” gene (necessary for non-sexual gene transfer)
◦ It leads to the dissemination and evolution of the genes.
Non-conjugative Plasmid:
◦ Not capable of horizontal gene transmission.
◦ Transmission takes place by the means of conjugative plasmid.
Intermediate Plasmid:
◦ Mobilizable.
◦ Carry only a single subset of gene.
◦ Increase the frequency of transferring of conjugative plasmids after parasitizing.

6. Cont…
On The Basis Of Their Function
a. F-plasmid.
b. R-plasmid.
c. Col-Plasmid.
d. Vir-plasmid.
e. Degradative plasmid.

7. F-plasmid
•Fertility plasmid.
•Self transmissible.
•An episome
•Large plasmid, allow gene transfer between bacteria.
•Capable of expression in Sex pilus.
•Not a treat
e.g. Escherichia coli etc.
DNA transfer
element

8. R-plasmid
Resistance plasmid.
Contain genes which are necessary for antibiotic
resistance.
Encodes enzymes which are capable of modifying or
destroying the target antibiotics.
Some have single antibiotic resistance gene whereas
some have many other antibiotic resistance genes.
Most are Mobilizable.
Unable to encode sex pilus.
Spreading is real threat.
e.g. Salmonella enteritidis etc

9. Col-plasmid
col-colicine bacteriocines
• Col plasmids are present in different genes of E.coli.
• They contain genes that control the synthesis of proteins called Colicines/bacteriocin.
(proteins which has the ability to kill other bacterial strains and are often used by
host bacterium).
• This colicines inhibit growth of related bacteria that lacks Col plasmid.
• Different types of colicines exhibit different mode of action.
• Col-B induces damage of cytoplasmic membrane of the target bacteria.
• Example of Col plasmids- (Col E2 and Col E3) causes degradation of nucleic acids.
• Bacteriocins frequently destroy cells by making channels in the plasma membrane by
this way expanding its permeability. They may sometimes also corrupt DNA or RNA or
act on peptidoglycan and weaken the cell wall.
• Col plasmids are may be self transmissible or non-self transmissible (this non self
transmissible may be mobilized by F plasmids).
• This means that when F+ cell contains Col E plasmid, this plasmid can integrate with
F factor and gets transported to F– cell during conjugation.

10. Virulence-plasmid
•A virulence plasmid is inside a bacterium, it transforms that
bacterium into a pathogen.
•In contrast with different non-virulent, bacteria that are pathogenic
in nature convey genes for virulence factors that permit them to
attack and infect their particular hosts.
•virulence plasmids can likewise be transmitted starting with one
bacterium then into the next.
•Other than virulence gene, plasmids have also been able to convey
other significant components that improve transmission and
maintenance of the bacterial cell.
•virulence plasmid can empower the organism to communicate a
variety of related virulence-associated functions providing the
organism with characters advantageous to survive in environment.
• They make the bacterium increasingly pathogenic as the bacterium
is better ready to oppose host defense or to produce toxins.

11. Degradative plasmids
•Degradative plasmids are Plasmids that encode genes required for the metabolism of wide
range environmental contaminants.
•As they can be transferred between microorganisms, they can provide a means for the rapid
horizontal spread of degradative genes among natural microbial populations.
•Direct seeding of plasmids by Soil bioremediation by borne genes into native soil is a potential
useful way to enhance the degradation of environmental pollutants.
•2-4-D plasmids were found in strains isolated by enrichment on 2-4-D as the sole source of carbon
and energy and some of them were found to degrade herbicide with similar structure.
•Strain of Pseudomonas Putida called NCIB was formed to possess plasmid PDTG1 with 83,042
base pairs. This plasmid also encodes enzymes for Naphthalene degradation.

12. Plasmids as vector
•Plasmids are generally composed of circular double chains of DNA. The two ends of plasmids are held together
by covalent bonds.
•Origin of Replication (ori): it refers to the site at which replication begins. In plasmids, this ori is
generally composed of A-T base pairs, which are much easier to separate during replication. As plasmids are
smaller in size, they have one to few origins of replication sites. Regulatory elements are also present at the ori
site. For example- Rep proteins.
•Multiple cloning sites: this is also called as polylinker. A short DNA sequence consists of few sites for
cleavage by restriction enzymes. At the cleavage site, strand can be cut by different polylinkers. One main
advantage of multiple cloning sites in plasmids is that it does not hinder the rest of the plasmid during the
process and also possess unique restriction enzymes, which can cut the plasmid at specific points to allow
DNA insertion.
•Antibiotic Resistance gene: This is one of the main components in plasmids which help in Drug resistance.
By a process of conjugation, plasmids transfer from one bacteria to the other and during this process they are
capable of conferring antibiotic resistance properties to the bacteria.
•A Promoter region: this region helps in the process of transcription and in recruitment of transcriptional
machinery.
•Primer binding site: this is specifically used for PCR amplification or for DNA sequencing and generally refers
to short sequence of DNA on a single strand.

13. A plasmids Vector