Intro to Y12 2025 Flashcards
Outline the steps (1-6) in bacterial transformation
Isolate gene for insulin chain A
Use reverse transcriptase to get c.DNA from isolated mRNA so that it contains no introns.
Cut gene and plasmid using the same endonuclease.
Stick insulin gene into plasmid using DNA ligase which joins the sugar phosphate backbone.
Transform bacteria using heat or electroporation and check for successful transformation by growing on an antibiotic plate with ampicillin and then an antibiotic plate with tetracycline
Bacteria with no plasmid will not grow on the ampicillin plate, successfully transformed bacteria with the non-recombinant plasmid will grow on both plates and the successfully transformed bacteria with the recombinant plasmid will only grow on the ampicillin plate.
How many polypeptides make up insulin?
2
What is the function of reverse transcriptase?
Convert c.DNA from isolated mRNA so that it contains no introns.
Why is reverse transcriptase used in the process?
So the introns are removed and a functional insulin is produced.
What is used to cut the plasmid when producing insulin?
Endonucleases
What is used to join the plasmid and insulin gene?
DNA ligase
What is the purpose of the AmpR and TetR genes in the plasmid?
To check for successful transformation of the bacteria
What is a recombinant plasmid?
A plasmid with the target gene (insulin gene)
What is the function of the promoter region?
Allows RNA polymerase to attach and transcribe the DNA
Which bacteria will only grow on ampicillin?
Bacteria that are successfully transformed with the recombinant plasmid
Which bacteria will grow on ampicillin and tetracycline?
Bacteria that are successfully transformed with the non-recombinant plasmid
Which bacteria will not grow on ampicillin or tetracycline?
Bacteria that have not been successfully transformed
Which antibiotics will bacteria that have been successfully transformed with the non-recombinant plasmid grow on?
Ampicillin and Tetracycline
Which antibiotics will bacteria that have been successfully transformed with the recombinant plasmid grow on?
Ampicillin only
Which antibiotics will bacteria that have not been successfully transformed with the recombinant plasmid grow on?
None (not ampicillin or tetracycline)
Qualitative data
Qualitative data is descriptions e.g. colour and is represented with a bar graph
Quantitative data
Quantitative data is numerical data e.g. 10 seconds and is represented as a line graph or histogram
Independent Variable
The independent variable is a factor that is changed or manipulated in an experiment to determine its effect on the dependent variable.
Dependent Variable
The dependent variable is measured to determine if the change or manipulation of the independent variable had any effect.
Control Variable
Something that is kept constant throughout the experiment.
Control group
A control group is where the factor affecting change is removed to provide a baseline for a comparison against the experimental group.
Experimental method scaffold
There will be two ….
In the experimental group………… will be changed by ………………….
Add …… to a test tube.
Add…..
Add….
The ……………..will be measured by ……………………after
All other variables will be kept the same e.g. ………………………
Repeat the above for the other ….*not relevant if it is an all or nothing.
For the control group repeat the above but with no ……………….
Repeat the experiment ………….
If ……………. then …………………
Accuracy
The accuracy of a measurement relates to how close it is to the ‘true’ value of the quantity being measured.
Improve accuracy
Accuracy can be improved by using a better measuring tool that is being used to measure the dependent variable or collecting quantitative data instead of qualitative data