Difference between revisions of "GDSYZX-United/PROJECT/extract dna"
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Revision as of 05:45, 16 October 2016
Extract DNA in arabidopsis thaliana
Place:Biology Experiment Teaching Center Of Sun Yat-sen University
Date:July 25th, 2016
Experimental apparatus:Grinding rods, 2 ml centrifuge tube, double panel, transfer liquid gun, adsorption column &collection tube and centrifuge.
Experimental drugs:Buffer PCB, β- mercaptoethanol, RNaseA, chloroform, Buffer BD, anhydrous ethanol, PW Solution, Wash Solution, TE Buffer.
Procedure:
| Experimental drugs | operation | Time |
|---|---|---|
| 50-100mg Young leaves of Arabidopsis thaliana. | Transfer to 2 ml centrifuge tube after grinding with grinding bar. | |
| 600μl 65℃ Buffer PCB & 12μl β- mercaptoethanol | Placed in 65 ℃ water after blending. | 25min |
| 20μl RNaseA | waiting | 5min |
| About 630μl chloroform | Transfer supernatant liquid to a clean 2 ml centrifuge tube after blending and following 12000 RPM centrifuge. | centrifuge：5min |
| About 400μl Buffer BD | Blending for 3-5 times. | |
| About 400μl anhydrous ethanol | Transfer to the adsorption column after blending,and then waiting. Dump the waste liquid after centrifuging with 10000 RPM. | waiting：2min centrifuging：1min |
| 500μl Wash Solution | Centrifuging with 10000 RPM after adding the reagent, and then dump the waste liquid. | 1min |
| centrifuge with 12000 RPM. | 2min | |
| 50μl TE Buffer | Place the adsorption column in the new 2 ml centrifuge tube, then waiting after adding reagent. | wait:3min |
| centrifuge with 12000 RPM. | 2min |
Result:
The target DNA can be extracted successfully in most of the groups, but they have very low concentration and many impurities.
Introspection:
- Understanding of the experimental requirements was not very well and had some wrong operations in the experiment.
- There is still many liquid residue on the grinding rod after grinding Arabidopsis thaliana leaves.
- Not all reagents were fully blended.
- The usage of transfer liquid gun was not normative.
arabidopsis thaliana
grinding with grinding bar
After blending with Buffer PCB
centrifuge
result(we falied this time)
