Test 2 - Cell culture and Gene transfection Flashcards
1
Q
Primary cell culture
A
The first culture grown in a dish from cells or tissue from the live organism.
2
Q
Secondary cell culture
A
Cell culture that is taken from a primary cell culture.
3
Q
Adherent Culture:
A
contact inhibition Regulatory mechanism that keeps cells growing in a single layer.
4
Q
Floating culture
A
In solution, does not require attachment to grow.
5
Q
Maintain cells?
A
- Immortal cells obtained two ways
- Gene transfection
- Simian Virus 40
- Over expression of TERT
- Random mutation providing oncological traits
- Freezing cells for later use.
- Gene transfection
6
Q
Viral vector
A
Does not need a mediator
7
Q
Non viral vector:
A
Plasmid DNA + Mediator
Can be delivered as naked DNA.
8
Q
Transient Transfection?
A
- DNA not incorporated into cell nucleus
- Used to measure promotor and enhancer activity or short term expression.
9
Q
Stable Transfection
A
- DNA incorporated into Cell nucleus
- Contains a antibiotic resistant marker that will protect the cell from the antibiotic used to kill any cells that didnt get the DNA.
- This is used for long-term studies of effects of protein overexpression, protein localization.
10
Q
How to remedy varied expression of cloned Gene?
A
- Perform single cell cloning, which will produce a stable cell line with a homogenous population:
- Extreme dilution
- Spread them out enough so you really only get one.
- Using cloning rings or cylinders
- Separates them so you know what you get.
- Extreme dilution
11
Q
What is Luciferase?
A
- Using a plasmid vector, insert the targeting DNA segments upstream of a luciferase gene.
- In the plasmid, there is no promoter or enhancer for this gene.
- Therefore, any luciferase activity will be the result from the inserted DNA segment.
