1. Molecular Biology Part I: Restriction Enzymes AP Lab 6

2. What you need to know! http://www.phschool.com/science/biol ogy_place/labbench/lab6/intro.htmlhttp://www.phschool.com/science/biol ogy_place/labbench/lab6/intro.html

3. Bacteria Contain: 1 chromosome 1 or more plasmids Restriction enzymes

4. Plasmids Small, circular DNA pieces Contain random DNA fragments that are collected or exchanged w/ other bacteria Contain nonsense information Sometimes contain useful information like antibiotic resistance

5. Restriction Enzymes (RE) Enzymes that cut DNA at very specific base sequences (often palindromes) Make blunt or sticky ends Evolved to combat invasive DNA from viruses Does not cut bacterium’s DNA because it’s missing correct DNA sequence Different bacterial strains have different RE

6. RE Nomenclature Named after the bacteria it comes from First capital letter is of the genus Lower case letters are the species Next capital letter is the strain The number is the order of discovery within the particular bacteria Example: EcoRI E = Escherichia co = coli R = RY13

7. EcoRI Cleaves double stranded DNA at GAATTC Cuts between the G and A on the top and bottom strand G|A A T T C C T T A A|G This creates a sticky end Other RE can create blunt ends –i.e.C C|G G G G|C C

8. Genetic Engineering RE fragments can be “glued” back together using ligase Biotech supply vendors offer: Purified RE in a large variety Plasmids with published restriction maps