Do all bacteria contain plasmids?

Introduction

During the 1940s when scientists found non-chromosomal DNA that’s responsible for antibody resistance, they called it by several names. Episomes, plasmagenes, plastogenes and bioblasts were some of the terms scientists called them.

In 1952, Nobel laureate Joshua Lederberg was the first to coin the term ‘plasmid’ by combining cytoplasm and ‘id’. Plasmids are mostly found in bacteria and some eukaryotes (multicell organisms).

What are Plasmids?

Plasmids are circular, small DNA pieces present in bacteria that can replicate without the influence of chromosomal DNA. While the genome contains millions or even billions of DNA base pairs, plasmids contain only a few thousands. Plasmids are small in size, relatively stable when compared to other DNA and can be easily replicated.

The presence of plasmids makes bacteria resistant to antibiotics and other poisons and helps them survive harsh conditions. Plasmids help life science engineers and biotechnologists in many ways. Lab-created plasmids are called constructs or vectors.

Parts of Plasmids

The origin of replication tells the plasmid the site from where to start the replication process.

• Antibiotic resistance genes occurring naturally in plasmids act to the bacteria’s advantage by providing resistance to antibiotics. When studied in a lab, this gene helps scientists identify bacteria that contain this gene from other cells that do not.

• With the help of a selectable marker, the DNA can be easily engineered by introducing foreign DNA into the plasmid sequence.

• The promotor site is where the RNA polymerase attaches itself to the gene and creates mRNA by transcription.

• Restriction sites are versatile regions in the plasmids which allows any genes to be engineered the way scientists require.

How genes are expressed?

The multiple cloning site is located at the restriction site in the DNA in the way of short segments which are flexible to allow cloning process.

The enzymes at the restriction site allow genes in that particular area to be cut using molecular scissors. Scientists can remove the original gene, and replace it with the gene of their choice and with the help of the promotor site, the gene sequence can be expressed by transcription to produce mRNA. Now, the gene sequence can be cropped to use somewhere else.

Genes, thus created, in a lab can then be used in several ways. This cropped gene can be inserted into other bacteria to study their properties. Genes produced in this way could be used to track a specific group of bacteria in a population.

Horizontal Gene Transfer

The movement of genetic code from one organism to the other is called horizontal gene transfer (HGT). This process occurs naturally and it is also responsible for the virulence and the amount of resistance a bacterium shows against antibiotics. There are four distinct ways in which a different DNA or RNA can be inserted inside a bacterial cell or into the cell of a eukaryotic organism namely transformation, transduction, conjugation and transfection.

While the first three types of DNA introduction into bacterial cells happen by nature using HGT, the latter (transfection) can be done only in a lab.

Transformation

When the plasma (outer) membrane of a bacterium is damaged or affected by external conditions, the plasmids tend to leak out of the cell wall making it easily available outside the lysed bacterium. When an adjacent bacterium takes in a plasmid from the environment it is called transformation. The presence of particular substances or different phases of cell growth naturally induces a transformation process in bacteria.

Not all bacteria’s cell walls are permeable enough for transformation to happen; thus, they are not competent enough to take in free plasmids. To make its cell wall permeable enough, scientists use calcium chloride to help the bacteria gulp the plasmids. Other methods such as the application of electric charge and electroporation are used to increase the permeability of the cell wall and therefore increase the efficiency of transformation.

Transduction

Using a virus or a viral vector, when a foreign DNA/RNA is introduced into a bacterium, it is called transduction.

Those viruses that can reproduce itself only inside a bacteria are called bacteriophages or phages. Such bacteriophages stick onto the membrane of the bacteria and inject the genetic material into it. Once the genetic material is injected, it can carry out any one of these life cycles inside the bacterium– lysogenic or lytic.

During the lytic cycle, the viral genome takes control of the bacteria’s host system and produces copies of its own. The bacterial cell lyses release copies of viral material to affect other bacteria.

During the lysogenic cycle, the viral genome attaches itself to the bacteria’s genome using a specific site of integration. Now, the phage will have to wait until the lytic cycle is triggered to carry out the process.

Both these life cycles pave way for the bacterial DNA unknowingly transferred into the freshly produced bacteriophages (viruses). When such copied DNA is transferred to other cells it is called transduction. Inside the infected bacterium, this transferred DNA can exist either in the form of a plasmid (outside the chromosome) or it can lie in an integrated form in the bacteria’s host genome as homologous or site-directed recombination.

Scientists use phagemids which contain the properties of both bacteriophages and plasmids to carry out transduction to introduce DNA sequences into a bacterium.

Conjugation

By direct contact when genetic code is transferred to a recipient bacterium from a donor, it is called conjugation. The fertility factor (F-factor) is a DNA sequence present in the donor bacterium. Episomes are long pieces of DNA sequence that can attach to the bacterial chromosome and replicates themselves in sync with it as long as the mother strain remains.

The F-factor present in the episome develops a small bridge-like structure called the sex pilus to draw the recipient bacterium closer. Once contact is made, the plasmids are transferred to the recipient.

Transfection

Transfection can only be done in a lab and only on eukaryotic cells, unlike the other three processes, using physical and chemical methods. Through non-viral methods, transfection is carried out by introducing foreign DNA into a eukaryotic cell. DNA molecules are either neutralized or a positive charge is imparted using chemicals like calcium phosphate and diethylaminoethyl (DEAE) – dextra to enable them to move through a negatively charged cell wall. Refer to figure 1.6. Electroporation involves creating transient pores using electric pulses in the cell membrane to let the DNA pass. Using a fine needle when genetic material is injected into the cell membrane, is called microinjection.

Plasmids are widely used to manipulate genes because of its versatility, cost-effectiveness, flexibility and safety. Bacteria can reproduce faster, and plasmids inside it can easily be grown, modified and used for a variety of purposes.

FAQs

Qns 1. What is Horizontal Gene Transfer (HGT)?

Ans. The transfer of genetic code from one organism to the other is called horizontal gene transfer (HGT). This process occurs naturally and it is also responsible for the virulence and the amount of resistance a bacterium shows against antibiotics.

Qns 2. What is transformation?

Ans. When the plasma (outer) membrane of a bacterium is damaged or affected by external conditions, the plasmids tend to leak out of the cell wall making it easily available outside the lysed bacterium. When an adjacent bacterium takes in a plasmid from the environment it is called transformation.

Qns 3. What is transduction?

Ans. Using a virus or a viral vector, when a foreign DNA/RNA is introduced into a bacterium, it is called transduction.

Qns 4. What is conjugation?

Ans. By direct contact when genetic material is passed on to a recipient bacterium from a donor, it is called conjugation.

Qns 5. What is transfection and where it can be carried on?

Ans. Through non-viral methods, transfection is carried out by introducing foreign DNA into a eukaryotic cell. Transfection can only be done in a lab and can be carried out only on eukaryotic cells.

Qns 6. What is mean by electroporation and microinjection?

Ans. DNA molecules are either neutralized or a positive charge is imparted using chemicals like calcium phosphate and diethylaminoethyl (DEAE) – dextra to enable them to move through a negatively charged cell wall. Electroporation involves creating transient pores using electric pulses in the cell membrane to let the DNA pass. Using a fine needle when genetic material is injected into the cell membrane, is called microinjection.