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Flashcards in Isolation Of Human Genomic DNA Deck (13):
0

What needs to be considered when choosing a DNA isolation method.

Cost effectiveness
Safety
Rapidity
Reliability
Yield
Purity

1

Main steps in DNA isolation

Cell lysis
Removal of proteins
Precipitation and cleaning of DNA

2

Role of sucrose

Isotonic sucrose preserves the integrity if WBCs before rupture
Allows washing away of broken RBCs before WBC lysis

3

Role of Triton X-100

Destroys membranes

4

Role of Tris Cl

Permeabilizes cell membranes by interacting with polysaccharide component

5

Role of EDTA

Binds divalent ions and inhibits DNAses

6

Role of SDS

Solubilise a proteins and lipids

7

Role of proteinase K

Removes protein

8

How to assess DNA integrity

- spectrophotometer
- gel electrophoresis

9

RFLP

Restriction fragment length polymorphism

10

How to figure out DNA concentration

Conc(ug/ml) = (A260 reading - A230 reading)x dilution factor x 50ug/ml

11

How to figure out total DNA yield

= DNA conc x total purified sample volume (ml)

12

Uses of isolated genomic DNA

- prep of genomic libraries
- gene/DNA sequencing
- analysis of genomic organisation
- DNA fingerprinting
- detection of abnormalities/mutations
- crime investigation
- paternity testing