Plasmid identification
- To prove that plasmid isolated by miniprep is pkd46
- Find out why plasmid isolated by miniprep appears to be so big in size

1. Found restriction sites on pkd46 by using some program on JustBio.com
2. Identify enzymes which cut plasmid at 1 point and those which cut it at 2 points
3. Check out which of those enzymes are available (Sal1 cut once to get a 6.3kb linear DNA, EcoR1 to cut twice, resulting in a 1.5kb fragment and a 4.8kb fragment)
4. Check out the buffer which the enzyme needs, the incubation temperature, inhibition method from the catalogs
5. Since we want to make 20ul, add
10x buffer 2ul
H2O 0ul
DNA(plasmid) 17ul
RE enzyme 1ul
6. Always add buffer and water first, then DNA, add enzyme last
7. Always keep DNA and enzyme in ice.
8. Incubate tubes in water bath at 37degrees for 2h
9. Inactivate enzymes at 65degrees for 20min
10. Store at -20degrees
11. Run gel tomorrow

Plasmid loss monitoring
DH5a NS3
Aim: To find out how much plasmid is lost during cell growth in selection and non-selective media
1. From LB+amp(?) plates, inoculate same colony into 2 LB flasks, 2 LB+amp flasks
2. Incubate at 37degrees, 24hrs
Will blog about the whole procedure when I do it tomorrow, a lot of stuff to do..