Transformation lab Flashcards
What is the purpose of the “pGlo plasmid” in the transformation experiment?
It acts as a vector to transport genes from one cell to another
Which statements are correct descriptions of the transformation solution? (select all that apply)
Helps to make the plasmid competent
contains calcium chloride
contains the plasmid
Provides nutrients for the transformed E. coli to grow
Helps to make the E. coli competent
- Contains calcium chloride
- Helps to make the E. coli competent
The E. coli in the “starter plate” is best described as:
The recipient cells
What is the purpose of LB broth?
Provides nutrients for the transformed cells to grow
A “lawn” means
bacterial growth is spread all over the surface of the medium
Which plate would you expect to have a lawn of growth, if the experiment goes as planned?
LB - DNA;
The LB plate has nothing in it to prevent every cell placed there from growing
What two things would have to happen in order for a cell to grow into a fluorescent colony?
- Transformation
- Taking up arabinose
The cell would have to be transformed in order to obtain the gene for florescence. Then, the gene needs to be “turned on” by the arabinose
Which plate would you expect to have fluorescent colonies?
LB/amp/ara +DNA;
Arabinose induces GFP expression in transformed cells
Which plate would you expect NOT to have any growth?
LB/amp -DNA;
Since LB/amp -DNA does not have transformed cells, no growth is expected because the untransformed cells lack ampicillin resistance
Think about experimental design for a moment. A well-designed experiment should have a control and an experimental aspect. In this experiment which are the control plates?
LB/amp -DNA: because it makes sure the antibiotic is working
LB/amp +DNA: because it tells you that the bacteria picked up the resistance gene (plasmid) transformation
(Question: Is E.coli able to pick up the plasmid?)
LB - DNA: because bacteria should grow on it under normal conditions, whether bacteria is alive or not
If there is no growth in the LB - DNA plate, what might explain this? (select all that apply)
The experiment worked as it should.
Transformation didn’t work.
The plate might have been mislabeled and it really contains ampicillin.
The plate could have been mislabeled and it really contains arabinose.
The student might have forgotten to inoculate the plate with bacteria.
The bacterial cells are not alive.
There is “something” in the LB medium that prevents “wildtype” E. coli from growing.
The plate might have been mislabeled and it really contains ampicillin.
The student might have forgotten to inoculate the plate with bacteria.
The bacterial cells are not alive.
There is “something” in the LB medium that prevents “wildtype” E. coli from growing.
What is the purpose of having the LB/amp -DNA plate in the experiment?
It tells us if the ampicillin works;
If untransformed cells grow on this plate, we know that the ampicillin isn’t working properly
Which two plates would you compare to see whether transformation occurred as planned?
LB/amp +DNA and LB/amp-DNA;
Comparing these two plates show the effect of transformation. The difference between these two is one has transformed cells (+DNA) while the other doesn’t (-DNA)
Which two plates would you compare to confirm the role that arabinose plays in gene expression?
LB/amp +DNA and LB/amp/ara +DNA;
These two plates are the same except that one has arabinose added to it. You should be able to see that arabinose “turns on” the GFP gene in the cells on the LB/amp/ara plate while the absence of arabinose shows no florescent cells on the LB/amp plate
A group of students carried out the transformation experiment. Their “LB/amp +DNA” plate has no growth. Their “LB/amp -DNA” plate contains colonies. The other two plates had the expected results. What is the MOST LIKELY explanation for this unusual result?
They mislabeled the two LB/amp plates
transformed (grow) vs untransformed (no grow)
