18.1: Introduction
Recombinant DNA Technology & Bacterial Transformation
In addition to the normal circular chromosome found in bacterial cells, some bacteria have a small extrachromosomal DNA molecule known as a plasmid . Plasmids replicate independently of the main chromosome and are not required for survival, growth, or reproduction. However, plasmids can confer survival advantages. Certain plasmids, for example, called R-plasmids , carry genes for resistance against antibiotics such as penicillin, tetracycline, or ampicillin. These genes encode enzymes that inactivate the antibiotic, allowing the bacterial cells to survive and grow.
In lecture, we introduced Frederick Griffith’s experiment on transformation. Bacterial transformation involves the transfer of foreign genetic information into bacterial cells. Transformed cells acquire new traits when they incorporate and express the genes found on the newly acquired DNA. In the laboratory, the type of DNA used most frequently for transformation experiments is bacterial plasmid DNA. The presence of antibiotic-resistance genes on plasmids makes it possible for scientists to select for bacteria containing the plasmid of interest. Bacteria that contain the plasmid will be resistant and able to grow in the presence of the antibiotic, whereas bacteria lacking the plasmid will be sensitive to the antibiotic and die.
Some bacteria, known as competent bacteria , are naturally receptive to foreign DNA. For other cells, competency can be generated artificially by changing the temperature and/or salt concentration of the medium. In today’s exercise, we will induce competent ampicillin-sensitive Escherichia coli cells to take up the plasmid pAMP, which contains a gene for ampicillin resistance. We will be able to select for transformed cells because only cells that have successfully taken up the plasmid will be able to grow on agar plates containing ampicillin. Any E. coli that haven’t been transformed will be killed by the ampicillin.
Figure \(\PageIndex{1}\): Predict the activity of competent, ampicillin-sensitive Escherichia coli cells when induced to take up the plasmid pAMP. Original image created by M. Chen.
Create a hypothesis and prediction for this experiment :
Hypothesis:
Prediction: