Question

Describe the mechanism of transformation.

Short answer

Answer: The key steps involved in the bacterial transformation process are 1) preparation of competent cells, 2) uptake of foreign DNA, 3) recombination of foreign DNA with the host genome, 4) expression of foreign DNA, and 5) selection and verification of transformed cells.

Step-by-step solution

Preparation of Competent Cells

To begin the process of transformation, bacteria must be made competent, meaning they are able to take up foreign DNA. This can be induced under laboratory conditions through chemical treatments, such as exposure to calcium chloride, or through applying an electric field (electroporation). These treatments increase the permeability of the bacterial cell membrane, allowing the foreign DNA to enter the cell.

Uptake of Foreign DNA

After preparing the bacterial cells, they are exposed to foreign DNA, usually in the form of a circular plasmid. The uptake of DNA occurs through the bacterial cell membrane, which has been made permeable by the treatments in Step 1. The foreign DNA binds to receptors on the cell membrane, and one strand of the DNA is then transferred into the cell through the membrane. The other strand is degraded by extracellular nucleases.

Recombination

Once inside the bacterial cell, the foreign DNA can recombine with the host cell's genomic DNA via homologous recombination. This is mediated by enzymes called recombinases, such as RecA in Escherichia coli. The process of recombination involves the exchange of homologous DNA regions between the foreign and genomic DNA, resulting in the integration of the foreign DNA into the host genome.

Expression of Foreign DNA

After successful integration into the host genome, the foreign DNA can be expressed by the bacterial cell. Transcription and translation machinery of the cell produce proteins encoded by the foreign DNA. In the case of transformation in a laboratory setting, the foreign DNA often includes a reporter gene or a selectable marker, which allows for the identification and selection of transformed cells.

Selection and Verification

Finally, the transformed bacterial cells must be distinguished from non-transformed cells. This is typically accomplished by including a selectable marker in the foreign DNA, such as an antibiotic resistance gene. When the bacterial cells are exposed to the corresponding antibiotic, only the transformed cells (carrying the resistance gene) will survive, allowing for their selection and further analysis to confirm the successful integration of the foreign DNA. In summary, the mechanism of transformation involves the preparation of competent bacterial cells, uptake of foreign DNA, recombination and incorporation of the foreign DNA into the host genome, expression of the foreign DNA, and selection and verification of transformed cells.