biotechnology – manipulation of biological organisms
(usually with DNA itself)
To study the functions of individual genes, molecular
biologists will cut them out of a genome and place
them into bacteria.
Six basic techniques will be covered:
restriction enzyme digest
Before DNA can be manipulated, it needs to be
isolated from the cells.
1. Cell membranes are disrupted
use a detergent
1. DNA precipitation
ethanol used to dehydrate and aggregate DNA
1. DNA isolation / storage
DNA must be cut into smaller pieces before they can
be used in other techniques.
restriction endonucleases – enzymes that cut DNA
backbones at specific sequences through hydrolysis
recognition site – the DNA sequence which restriction
enzymes bind to
unsure if enzymes scan DNA to find sequences
Why are these enzymes used?
These enzymes are naturally found in bacteria.
Bacteria found to be resistant to some bacteriophage
Restriction enzymes would cut viral DNA, not its own
Restriction enzymes cut at recognition sites.
Recognition sites are typically 4 to 8 bp in length.
They are always palindromic.
5’ G A A T T C 3’
3’ C T T A A G 5’
This sequence is specific for the EcoRI enzyme.
Restriction enzymes are named according to the
organism from which it was identified.
- strain RY13
- 1st enzyme in this strain