Plasmids are small, circular pieces of extrachromosomal DNA found in bacteria and other organisms. They can replicate independently of the chromosomal DNA and often contain genes that provide advantages to the host cell like antibiotic resistance. Key features of plasmids include an origin of replication, antibiotic resistance genes, and mechanisms for horizontal transfer between organisms like conjugation. Plasmids are important tools in biotechnology as they can be used to clone genes and amplify DNA fragments.

1. plasmids

2. CONTENTS
INTRODUCTION
PROPERTIES
ELEMENTS OF PLASMIDS
PLASMID REPLICATION
MODE OF TRANSFER
CLASSIFICATION / TYPES OF PLASMID
PLASMID AND RECOMBINANT DNA TECHNOLOGY
USES
IMPORTANCE OF PLASMID
EXAMPLES

3. INTRODUCTION
• A plasmid is a small, circular, extrachromosomal double
stranded DNA that has the capacity to replicate
independently of the chromosomal DNA.
• Discovered by Laderberg in 1952.
• Plasmids are found naturally in many microorganisms
• It naturally occur in bacteria, but sometimes found in
eukaryotic organisms.
• When we think of plasmids, we probably also think of
bacteria, but plasmids are not restricted to bacteria. In
fact, most S. cerevisiae(eukaryote) strains carry a large
plasmid of 2 micron or 2µm

5. EPISOMES
• Plasmid is an
extrachromosomal DNA
molecule whereas
episomes is any kind of
extra-chromosomal
DNA that can link up
with chromosomal DNA.
• Eg of episomes are
viruses, because they
integrate their genetic
material into host’s
chromosomal DNA.

6. Defination
EPISOME: plasmid of bacteria
or viral DNA that can integrate
itself into the chromosomal
DNA of the host organism. For
this reason, it can stay intact for
long time, be duplicated with
every cell division of the host,
and become the part of its
genetic makeup. At some stage
may exist as independent
element.

7. PROPERTIES
 Size range from 1kbp to several mbp.
 Replicate independently and code for their own transfer.
 Do not cause damage to cells in which it harbour and generally
are beneficial, do not have extracellular forms and exist inside cells
simply as free and typically circular DNA
 Plasmids are non essential for existence of bacterial cells except
under specific environment.
 Plasmids contains genetic information for their own replication.
Genes carried by plasmid encodes trait for antibiotic resistance,
heavy metal resistance, nitrogen fixation, and pollutant
degradation.
 Some produces virulence factor that help in defence or nutrient
utilization.

8. Number of plasmids in an individual cell may vary,
ranging from one to several hundreds, denoted by
copy number.
 Stringent replication
Single copy plasmids
replicate and segregate with
the bacterial chromosome.
This is called as stringent
replication
 Relaxed replication
Multiple copy plasmids
undergo more than one
replication for each
replication of the bacterial
chromosome. This is called
relaxed replication.

9. Copy number
 Copy number is average or expected
number of copies per host cell.
Plasmids are either low, medium, or
high copy number.
 The Copy number refers to the
number of molecules of an
individual plasmid that are normally
found in a single bacterial cell.
 Larger plasmids have a low copy
number of just 1 or 2 per cell.
 Other called relaxed plasmids, are
present in multiple copies of 50 or
more per cell.
 A useful cloning vector needs to be
present in the cell in multiple copies
so that large quantity of
recombinant DNA WILL ONTAINED.

11. ELEMENTS OF PLASMIDS
 Origin of replication: it is the DNA sequence which directs
initiation of plasmid replication by recruiting bacterial
transcriptional machinery.
 Antibiotic resistence gene: these genes allows for selection
of plasmid containg bacteria by providing a survival
advantage to the bacterial host.
 Multiple cloning sites: this is the short segment containing
several restriction enzyme sites, enabling easy insertion of
foreign DNA.

12. ELEMENTS OF PLASMIDS
 Origin of replication: it is the DNA
sequence which directs initiation of
plasmid replication by recruiting
bacterial transcriptional machinery.
 Antibiotic resistance gene: these
genes allows for selection of
plasmid containing bacteria by
providing a survival advantage to
the bacterial host.
 Multiple cloning sites: this is the
short segment containing several
restriction enzyme sites, enabling
easy insertion of foreign DNA.

13. • Insert: it is the foreign DNA cloned into the
multiple cloning site.
• Promoter region: it drives the transcription of
foreign insert .
• Selectable marker: it is used to select for cells that
has successfully taken up the plasmid for the
purpose of expressing the insert.
• Primer binding site: it is the site for binding of
short single stranded DNA sequence, used as an
initiation point for PCR amplification or sequencing
of the plasmid.

14. PLASMID
CONFORMATIONS
(a) Nicked open circular DNA
(b) Relaxed circular DNA
(c) Linear DNA
(d) Supercoiled or covalently closed circular
DNA

16. PLASMID REPLICATION
Plasmids replicate autonomously because they have
their own replication origins.
 Most plasmids in gram-negative bacteria replicate in a
manner similar to the replication of bacterial
chromosome involving initiation at the replication origin
site and bidirectional replication around the DNA circle
giving a theta(ø) intermediate.
 Most plasmids of gram-positive bacteria replicate by a
rolling circle mechanism.

17. Theta model
A theta structure is an
intermediate structure
formed during the
replication of circular
DNA molecule
(prokaryotic DNA), two
replication forks can
proceed independently
around the DNA ring and
it resembles the greek
letter theta(ø)

18. Rolling circle mechanism
1. Cut at ‘ori’ site.
2. elongation: using the
unbroken strand as
template. Polymerase will
start to move in a circle for
elongation.
3. termination: linear DNA is
cleaved from circle, resulting
double stranded circular
DNA and a ss linear DNA.
Linear ss DNA is circularized
by the action of ligase and
then replication to form
double stranded circular
plasmid.

19. Continued..

20. MODE OF PLASMID TRANSFER
The genetic information encoded in a
plasmid of bacteria is transferred
acorss a broad range of
microorganisms via-
1. TRANSFORMATION: requires
competent cells which are ready
to accept extracellular plasmid
and further stable replication
inside host cell.
2. TRANSDUCTION: plasmid
mediated gene transfer through
bacteriophages.
3. CONJUGATION: transfer through
cell to cell contact of donor and
recipient cell, require DNA
metabolism of donor cell.

22. CLASSIFICATION
 Plasmids are classified depending on their different function in
a host.
 Depending on their replication property, broadly, they can be
classified as compatibility and incompatibility groups.
 When two different plasmids when two plasmids with
can coexist stably in a cell, similar replicon or
they are called compatibility replication element are
plasmid. Unable to coexist called
incompatibility of plasmid

23. Incompatibility of plasmids
• Plasmid incompatibility is failure of
2 plasmids to coexist in a single cell.
• COPY NO AND PLASMID
INCOMPATIBILITYARE
LINKED:
• The no of plasmid in a cell is
governed by elements encoded
within the ori. It is not possible to
maintain two different plasmids that
use the same mechanism for
replication in a single cell.
• It is widely accepted that
competition for replication factors
leads to competition b/w plasmids
When a cell contain 2 incompatible plasmids, cell cannot
distinguish between the origins at the point of replication
initiation. Resultant repli. is postponed until after the two
plasmids have been distributed to diff. cells to restore the pre-
replication copy number.

24. Depending on their property plasmids are classified as F plasmids, colicinogenic (Col)
plasmids and R plasmids.
1. F-plasmid: F (fertility) factor., these plasmids are responsible for conjugation and are
also called conjugative plasmid. Due to their presence bacteria cell behaves as male.
2. Col plasmid: these are the groups of small multicopy colicinogenic plasmids which
encode the genes to synthesize colicins (bacteriocins). These plasmids need DNA
polymerase I for replication. These plasmids are widely used in gene cloning process
and are also used a s a good model for gene replication, transcription, and
translation.
3. R-PLASMID: this plasmid carry genes for resistance to antibiotics., like
chloramphenicol, tetracycline, streptomycin, sulfisomidine, ampicillin, trimethoprim
Most of them belong to incompatibility group of plasmids.

25. There are several types of other bacterial plasmids like
suicide plasmids, virulence plasmids but the above
discussed plasmids are important and are widely
studied ., and are used in genetic engineering…………..

26. F-PLASMID (fertility
plasmid)
• Carries the fertility
genes for
conjugation, transfer
the genetic
information between
two cells.
• Contain ‘tra’
(transfer) gene
R-PLASMID(resistance
plasmid)
• Contains genes that
provide resistance
against antibiotics or
poisons.
Col plasmid
• Contains genes that
codes for
bacteriocins (protein
that can kill bacteria)

27. DEGRADATIVE
PLASMID
• they enable the
digestion of unusual
substances that not
commonly found in
nature like toluene
and salicylic acid.
VIRULENCE
PLASMID
• They turn the
bacteria into
pathogen
• Ti plasmids of
Agrobacterium
tumefaciens, induce
crown gall disease
on dicots plants
SUICIDE PLASMID
• Gets transferred to
another bacterial cell
but don’t replicate
further.

30. PLASMIDS AND RECOMBINANT DNA
TECHNOLOGY
• Recombination DNA technology is joining together of DNA molecules
from two different species that are inserted into a host organism to
produce new genetic combinations that are of value to science, medicine,
agriculture, and industry.
• Artificially constructed plasmids are used as vectors in genetic engineering
to clone or amplify or express particular gene
• When a single recombinant DNA molecule composed of vector +
inserted DNA is introduced into host cell, the inserted DNA is reproduced
along with the vector, producing large no. of recombinant DNA molecule.

33. USES
1. CLONING
2. PROTEIN PRODUCTION
3. GENE THERAPY
4. GENETIC DISEASE MODEL
5. PLASMID DNA VACCINE

34. Importance of plasmid
 Easy to work with due to convenient size for physical isolation and
manipulation
 Independent origin of replication allows plasmid replication in the cell
to proceed independently from the direct chromosomal control
 Multiple copy number makes them to be present in the cell in several
copies to that amplification of the plasmid DNA becomes easy.
 Presence of selectable markers such as antibiotic resistance genes,
which make detection and selection of plasmid-containg clones easier.

35. Continued…
 stable for long term either as purified DNA or within
bacterial cell preserved as glycerol stocks.
 Functional in many species including plants, worms, mice,
and even cultured human cells and useful for a diverse set
of applications like investigation of promotors , small RNAs
or other genetic element.
DRAWBACK: less useful for cloning large segment of
DNA(>10kbp).

36. EXAMPLES
• pBR322
• pBR327
• pBR325
• pBR328
• pUC8
• pUC9
• pUC12
• pUC13

37. References..
 Principles of molecular biology by VEER BALA RASTOGI.
 Banu and Prasad, (2017) “Role of Plasmids in Microbiology”. J Aquac Res
Development.
 T. A. Brown (2010). "Chapter 2 - Vectors for Gene Cloning: Plasmids and
Bacteriophages". Gene Cloning and DNA Analysis
 Molecular biology by clarks.

38. THANK YOU……