R
amp
then:
7. If there is successful rDNA formation with
A No growth on penicillin medium
B. Growth occurs on penicillin medium
C. Both a and b
D.None
Answers
Answer:
Explanation:
Information for Teachers
Safety Instructions
Although the E. coli strain used in these experiments has been rendered non-pathogenic, it is important to teach the students good sterile technique and safe disposal of bacteria.
Gloves and safety glasses are to be worn at all times during this experiment.
Keep nose and mouth away from tip end when pipetting suspension culture to avoid inhaling any aerosol that might be created.
Use a 10% bleach solution to wipe down the benches at the end of the experiment.
Wash hands before leaving lab.
To dispose of contaminated material:
Immerse all disposable pipets, tubes, and loops that have come in contact with bacteria in 10% bleach solution for at least 20 minutes before draining, rinsing and disposing of in the trash.
When lab is complete, collect all petri dishes, open, and immerse in a 10% bleach solution to kill all bacteria. Allow materials to stand in bleach solution for 20 minutes or more. Drain excess solution, seal materials in a plastic bag and dispose in the trash.
Introduction
Transformation of cells is a widely used and versatile tool in genetic engineering and is of critical importance in the development of molecular biology. The purpose of this technique is to introduce a foreign plasmid into bacteria, the bacteria then amplifies the plasmid, making large quantities of it.
A plasmid is a small circular piece of DNA (about 2,000 to 10,000 base pairs) that contains important genetic information for the growth of bacteria. Bacteria, which often grow in the same environment as molds and fungi, evolved to make proteins that inactivate the toxins produced by these other organisms. The bacteria share this vital information by passing it among themselves in the form of genes in plasmids. Hence, the natural function of a plasmid is to transfer genetic information vital to the survival of the bacteria. It is this characteristic of plasmids that is exploited for use in transformation.
For bacteria to take in a plasmid, they must first be made "competent" to take up DNA, which won't normally pass through a bacterial cell's membrane. This is done by creating small holes in the bacterial cells by suspending them in a solution with a high concentration of calcium. DNA can then be forced into the cells by the procedures followed during this experiment.
In this activity, students will use a strain of E. coli that has been made competent to allow it to incorporate and express a plasmid containing two genes. One gene codes for a green fluorescent protein (GFP) and the other codes for ampicillin resistance. The source of the GFP gene is the bioluminescent jellyfish Aequorea victoria. The ampicillin-resistance gene allows us to select which of the E. coli cells have been transformed based on their ability to grow in an environment that contains the antibiotic ampicillin.
Figure 01 - Click to Enlarge
Objectives
Understand recombinant DNA techniques, in particular the transformation procedure using the heat shock method.
Understand the uses of marker or reporter genes in molecular biology experiments and how to screen for a gene of interest.
Understand that DNA can be transferred to another organism and therefore change the observable characteristics of that organism.
Become familiar with sterile technique and decontamination procedures that are used to handle bacteria.
Learn how to calculate transformation efficiency.