Isolation Of Human Genomic DNA

Question 0

Main steps in DNA isolation

Answer 0

Cell lysis
Removal of proteins
Precipitation and cleaning of DNA

Question 1

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

Answer 1

Cost effectiveness
Safety
Rapidity
Reliability 
Yield
Purity

Question 2

Role of sucrose

Answer 2

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

Question 3

Role of Triton X-100

Answer 3

Destroys membranes

Question 4

Role of Tris Cl

Answer 4

Permeabilizes cell membranes by interacting with polysaccharide component

Question 5

Role of EDTA

Answer 5

Binds divalent ions and inhibits DNAses

Question 6

Role of SDS

Answer 6

Solubilise a proteins and lipids

Question 7

Role of proteinase K

Answer 7

Removes protein

Question 8

How to assess DNA integrity

Answer 8

• spectrophotometer

- gel electrophoresis

Question 9

RFLP

Answer 9

Restriction fragment length polymorphism

Question 10

How to figure out DNA concentration

Answer 10

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

Question 11

How to figure out total DNA yield

Answer 11

= DNA conc x total purified sample volume (ml)

Question 12

Uses of isolated genomic DNA

Answer 12

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