Microbiological context of Bacterial Plasmid

1. Bacterial Plasmid
Dr. Avizit Sarker
MBBS (DMC), MD (Microbiology), BCS (Health)
Department of Microbiology
Dhaka Medical College

2. History
• The term plasmid was first introduced by the American
molecular biologist Joshua Lederberg in 1952.
• He was an American molecular biologist known for his
work in genetics, artificial intelligence and space
exploration.
• He won the Nobel Prize in Physiology or Medicine in
1958 for discovering that bacteria can mate and
exchange genes. He shared the prize with Edward
Tatum and George Beadle who won for their work
with genetics.

3. Plasmid
• A plasmid is a small, circular, double-stranded
DNA molecule within a cell that is physically
separated from a chromosomal DNA and can
replicate independently.
• Plasmids are present in bacteria, archaea &
eukaryotic organisms.

4. Properties of Plasmid
1. Small, circular, double-stranded DNA molecule can exist
independently of host chromosomes.
2. They have their own replication origins and are autonomously
replicating stably inherited.
3. It has relatively few genes. The size of the plasmid varies from
1-10 Kilo-base pair (kbp).
4. Plasmid can be transmitted from one bacterium to another.
5. Can be essential for specific environments: virulence,
antibiotics resistance, use of unusual nutrients, production of
bacteriocins.

5. Structure of Plasmid
• Origin of replication (replicon) - The origin of replication (ori) refers to a
specific location in the strand at which replication begins. For plasmids,
this location is largely composed of A-T base pairs that are easier to
separate during replication.
• Antibiotic resistance gene - The antibiotic resistance gene is one of the
main components of plasmids. These genes play an important role in drug
resistance (to one or more antibiotics) thus making treatment of some
diseases more challenging.
• Multiple cloning sites - Short segment of DNA which contains several
restriction sites allowing for the easy insertion of DNA.
• Promoter region - This is the component of plasmids that is involved in
recruiting transcriptional machinery.

6. • Primer binding site - This is a short sequence of DNA on a single strand
that is typically used as an initiation point for the purposes of PCR
amplification or DNA sequencing.
• Insert - Gene, promoter or other DNA fragments cloned into the MCS for
further study.

7. Classification of Plasmid
A. Based on ability to perform conjugation
1. Conjugative plasmids
Conjugative plasmids contain a set of transfer or tra
genes which promote sexual conjugation between
different cells. In the complex process of conjugation,
plasmid may be transferred from one bacterium to
another via sex pili encoded by some of the tra genes.
2. Non-conjugative plasmids
Non-conjugative plasmids are incapable of initiating
conjugation, hence they can be transferred only with
the assistance of conjugative plasmids.

8. B. Based on compatibility
1. Compatible - Microbes can contains different types of plasmids if they are compatible
to each other.
2. Incompatible - Can not co-exists in a microorganism.

9. C. Based on function
1. Fertility or F-plasmids - Contains tra genes. They are capable of conjugation and express sex pili.
2. Resistance (R) plasmids -
Structure -
a) Resistance transfer factor : carrying all genes necessary for conjugal transfer.
b) r- determinants : containing all or most of the antibiotics resistance genes.
Properties of r factors -
• Responsible of drug resistance.
• They can be transferred not only to the cells of the same species but also to other
species.
• Resistance to metal ions.
• Resistance to certain bacterial virus by coding for restriction endonuclease that
degrade the DNA of the infecting bacteriophage.

10. 3. Col plasmids - contains genes that code for bacteriocins.
4. Degradative plasmids - they enable the host in various metabolic activities.
• Digestion of unusual substance, like toluene, salicylate etc.
• Urease synthesis.
• Nitrogen fixation.
5. Virulence plasmids - they code for certain virulence factors and toxins that
help in bacterial pathogenesis.
• Heat labile & heat stable toxin of E. coli.
• Siderophore production.
• Adherence antigens.

11. Bacterial cells depending on F factors :
• 1. F+ cell

12. • 2. Hfr cells (High frequency of recombination)
• Cells that contain F plasmid free in the cytoplasm (F+ cells) have no
unusual character apart from the ability to produce pili to transfer F to
F- cells by conjugation.
• In very small proportion of F+ cells, F factors become inserted into
bacterial chromosome.
• These type of cells are called Hfr cells.

13. • 3. F’(Prime) cell

14. Phenotypic effects of plasmids
A. Resistance effect
1. Resistance to antibiotic.
2. Resistant to heavy metal - Arsenic, Mercury, Cobalt etc.
3. Resistant to anion - Aresnite, Telurite.
4. Resistant to UV ray.
B. Metabolic
1. Antibiotic production by Streptomyces.
2. Metabolism of CHO, Protein, Gelatine, Casine.
3. Nitrogen fixation.
4. Citrate utilization.
5. H2S production.
6. Leucine biosynthesis.

15. C. Pathogenic effect
1. Antibiotic resistance & bacteriocin production
2. Toxin production
3. Colonization antigen
4. Tumorigenicity
D. Conjugal effect
1. Sex pili formation
2. Surface exclusion
3. Fertility inhibition

16. Application of Plasmid
Plasmid as a vector
• A vector is a DNA molecule used to carry genes from organism to organism.
Plasmid have an origin of replication, multiple cloning site, a selectable
marker gene and used in genetic engineering.
Protein production
• Human insulin production by recombinant DNA technology.
DNA vaccine
• Plasmid DNA is the best for promising DNA vaccine & gene therapies
against many infections, acquired & genetic disease including HIV-AIDS,
Malaria, different types of cancer, enteric pathogens & influenza.

18. Question
• 1. Classification of plasmid.
• 2. What is R factor? Properties of R factor.
• 3. Application of plasmid in genetic engineering.