Team:UT-Dallas/Project/methods

Methods and Protocols

Project

Miniprep

We used QIAGEN's miniprep reagents and protocol.

1. Pellet cells at 4000 rpm, 4C for 12 minutes
2. Resuspend cells with 200ul buffer P1, transfer to microcentrifuge
3. Add 200 ul lysis buffer P2, cap and shake. Do not allow reaction to continue for more than 5 minutes.
4. Add 300 ul Neutralization buffer N3, cap and shake.
5. Pellet cells at 13000 rpm for 10 minutes with tabletop centrifuge.
6. Apply supernatant to spin column.
7. Centrifuge for 60s. Discard flow through.
8. Add 750 ul wash buffer PE to column.
9. Centrifuge for one minute. Discard flow through.
10. Centrifuge again for one minute to get rid of excess wash buffer.
11. Transfer column to microcentrifuge tube. Add 50 ul elution solution EB to the column, let stand for 1-5 minutes.
12. Centrifuge for 60s. Determine concentration of DNA product and store at -20C

Gel Extraction

We used QIAGEN's Gell extraction protocol and reagents.

1. Use razor to excise gel slice containing desired DNA.
2. Slice this gel finely and place in a microcentrifuge tube.
3. Add 750 ul QG buffer to tube.
4. Incubate at 50C while shaking tubes at 500 rpm. You may need to vortex the tubes occasionally. Continue until the gell has completely dissolved. This is usually around 10 min.
5. Add something

Dinosaur

This is a dinosaur!!

Some Other Protocol

Well, aren't we interesting

More about our project:

• History of Cholera
• Project Details
• Materials and Methods
• The Experiments
• Results
• Data analysis
• Conclusions