PCR, Bacteriophages, and Bacterial Genetics: A Comprehensive Study

Posted on Jan 20, 2025 in Biology

PCR Technique

The Polymerase Chain Reaction (PCR) is a molecular biology technique that allows the amplification of a specific DNA fragment, generating a large number of copies. It is based on cycles of temperature increases and decreases that enable DNA denaturation and DNA polymerase activity, respectively. After PCR amplification, it is easier to identify viruses or bacteria. PCR has several applications:

Sequencing

PCR is responsible for generating sufficient DNA template for sequencing. It is easier and faster than cell cloning, and sequencing can be performed in a single operation.

Studies of Expression

PCR can be used in the study of mRNA produced by gene expression. RNA is reverse transcribed to give cDNA, which is then amplified. Studies can show when genes, including viral or tumor genes, are being expressed.

Evolutionary Biology and Mapping Studies

With its great ability to work with small amounts of low-quality DNA, the technique allows the analysis of tissues from extinct species. It can also be used to map the human genome.

Structure of Bacteriophage

Bacteriophages are a complex group of viruses composed of several parts of different shapes and symmetry. They consist of an icosahedral head, which houses the nucleic acid, and a helical tail that can inject the contents of the head. The head and tail are separated by a necklace made of capsomeres. At the end of the tail is the basal plate, which is fixed to the host cell. Several fibers that enhance fixation extend from the basal plate.

Lytic Cycle

The lytic cycle occurs when the genetic material of the virus enters the host cell and begins to make new viruses. The stages of the lytic cycle are:

  1. Adsorption Phase: The bacteriophage tail fixes to the wall of the bacteria.
  2. Penetration Stage: The tail contracts and injects the nucleic acid inside the bacteria.
  3. Lag Phase: The viral nucleic acid disrupts the functioning of the cell. From this point, the cell produces viral components.
  4. Assembly Phase: Viral components are assembled to originate new viruses.
  5. Lysis Phase: The bacterial wall is destroyed, and new viruses are released to infect new cells.

Differences with the Lysogenic Cycle

In the lysogenic cycle, those genes that allow the synthesis of viral proteins may remain suppressed indefinitely. The viral DNA is inserted into the DNA of the bacteria. These are called temperate, attenuated viruses, or prophages. This viral DNA can be passed to subsequent generations after bacterial reproduction. When conditions change (e.g., by ultraviolet radiation), the viruses enter the lytic cycle to reproduce new viruses until the cell bursts.

Asexual Reproduction and Parasexual Processes in Bacteria

Bacterial reproduction is primarily by asexual binary fission or bipartition. After DNA replication, regulated by the DNA polymerase of mesosomes, a transverse wall forms that will cause the formation of two daughter cells. In an environment with optimal conditions, bacteria can divide every few minutes.

Parasexual Processes

  • Transformation: This consists of the capture of DNA from another bacterium, which was free in the medium, and its incorporation into the bacterial chromosome. It does not involve any carrier agent.
  • Transduction: For the transfer of genetic material, the vector or carrier agent is a virus (phage) that acts as an intermediary between bacteria. The DNA does not need to be free in the medium.
  • Conjugation: This is when the transfer of DNA is carried out through the fimbria or pili from a donor bacterium to a recipient bacterium.

Bacterial Cell Wall Types

The bacterial cell wall is a rigid envelope surrounding the plasma membrane. According to the structure of the cell wall, two different groups can be identified using staining: Gram-positive (Gram+) and Gram-negative (Gram-). In both, the wall consists of a peptidoglycan called murein. The cell wall has several functions, such as giving the cell its shape, providing protection to avoid destruction by osmotic pressure, and housing most bacterial antigens.

Gram-Positive Bacteria

Gram-positive bacteria have thick, uniform walls without an outer membrane. Their walls are constituted by layers of peptidoglycan reinforced with teichoic acid. These bacteria are stained violet with Gram stain.

Gram-Negative Bacteria

Gram-negative bacteria have a thin and complex wall. They have a single inner peptidoglycan layer and an outer membrane consisting of a layer of phospholipids and another layer of glycolipids associated with polysaccharides that project to the outside. This membrane is permeable. These bacteria are stained red with Gram stain.