Genetic Engineering Flashcards
Are the three types of blotting done in situ or homogenate?
Homogenate
What is southern blotting used for visualising?
DNA
What is western blotting used for visualising?
Proteins
What is northern blotting used for visualising?
RNA
What is an in situ method for visualising RNA?
In situ hybridisation
What is a method to visualise protein in homogenate?
Western blotting
What is used to visualise DNA in situ?
Chromosome painting/Chromosomal spreads
What is used to visualise protein in situ?
Immunocytochemistry/Immunohistochemistry
What are the advantages of visualising in homogenate?
- Quantification
- Size
- Isolation
What are the disadvantages of visualising in homogenate?
- Requires more tissue
What are the advantages of visualising in situ?
- Tissue distribution
- Function
- Changes in above
What are the disadvantages of visualising in situ?
- Requires tissue processing
- Limited and compromised by reagents and resolution
3 Steps of visualisation and detection of DNA, RNA and protein in homogenates
- Gel electrophoresis
- Blotting (transfer out of gel into a nylon membrane)
- Detection by probing
What is a gel?
A gel is a porous matrix that acts like a sieve. Pores are present between the gel particles
Gel electrophoresis
DNA is loaded into a well on negative side
Electric current through gel and due to negatively charged phosphate moves towards positive electrode
Smaller molecules move further in the set time
Compare bands with DNA standard