Purpose: 

The purpose of this lab meeting was to purify the DNA extracted and make agarose gels for gel electrophoresis. This was done in order to prepare for DNA analysis using the gels to determine a possible match of DNA in the soil when comparing the standard ladder.

Procedure: 

DNA Purification: 

1. Before beginning make sure to obtain the proper PPE, since the lab deals with potentially harmful chemicals gloves should be worn at all times.
2. Use a microfuge tube with the crude soil and fill with sterile water until the 1 mL mark is reached.
3. Transfer solution to a 15 mL tube that can be used for mixing.
4. Add 2 mLs of warm DNA resin from the kit, this should be at 37ºC and mix by inversion several times before pipetting 2 mL to tube.
5. Set up column on syringe barrel, and set on vacuum filtration manifold.
6. Add roughly half of the resin to the column and urn on the vacuum. When the liquid is pulled through add the remainder of your sample.
7. Was column by adding 2 mL of 80% isopropanol and using the vacuum to pull through. Begin by adding small amount and allowing to flow before adding more.
   1. Repeat twice more for a total of three washes.
8. Remove column from barrel and put column in clean 1.5 mL tube. Spin at 8000 x g for 5 minutes in centrifuge to remove residual isopropanol.
9. Take column out of 1.5 mL tubes and put in 80ºC+ heat block for about thirty seconds to one minute.
10. Place column in new 1.5 mL tubes and apply 50 microliters of diH2O heated to 80ºc directly to column.
11. Incubate for one minute
12. Spin at 8000 x g for one minute

Making Agarose Gel

1. Transfer 4 mL of 10x stock TAE to 36 mL of diH2O in vile.
2. Add 0.4 g of agar powder to a 125 mL Erlenmeyer flask, add the TAE solution to the flask.
3. Place flask in microwave for one minute, once done mix the solution by swirling the flask.
4. Add 2 microliters of ethidium bromide to the flask.
5. Transfer the solution in the flask to to gel cast and let solidify (30 minutes).
6. Pour 1x TAE on top of the solidified gel until the cast is full.
7. Obtain a plastic bag (ziploc) and store in refrigerator at 4ºC for one week.

Data: 

No observations or data was recorded.

Conclusion: 

In this lab we purified the DNA to prepare it for gel electrophoresis.

Future Goals: 

I want to expand my knowledge on the processes and how everything actually works instead of just mixing a bunch of different chemicals together. Next lab meeting hopefully the gels will show successful results.