Basic Nucleic Acid Extraction Methods Flashcards
Beer-Lambert’s Law
A = log¹⁰ (I⁰/I) = ecl
Steps in DNA extraction
- Pretreatment and washing
- Cell lysis
- Removal of contaminants
- DNA precipitation
- DNA suspension
Physical methods on cell lysis:
Grinding
Shearing
Bead beating
Freeze-thaw
Types of solution-based method on cell lysis
Chemical
Enzymatic
What does SDS mean, and how does it remove contaminants in DNA extraction?
Sodium dodecyl sulfate
It makes the proteins anionic
A broad-spectrum serine protease that can also degrade nucleases
Proteinase K
What is used to remove RNA from a DNA extraction?
RNAse
Extraction methods on DNA precipitation
Organic
Inorganic
Solid phase
How does the organic method separate DNA from other components in a solution?
Uses high salt concentration in low pH with 1:1 phenol-chloroform
Solvation shell forms around the molecules
Inorganic method is also called as ______
Salting out
Optimal ratio of A260/A230 for DNA & RNA
2.0-2.2
Optimal ratio of A260/A280 for DNA & RNA
1.8-2.0
When dissociating a tissue, the sample must be _______ to inactivate intracellular RNAses
Frozen in liquid nitrogen or immeresed in a buffer solution