Week 3: Plasmid Extraction

 Introduction: 

    Since our plates and flasks had plenty of growth and no contamination, we were able to move forward into plasmid extraction using the GeneJET Plasmid Miniprep kit. 

Process: 

    Basically, we followed the protocol that was given to us from the kit. However, before starting the kit, we must extract the DNA of D. Aquaticus from the cells. We would use Eppendorf tubes to single out the bacteria pellet. We used 4 tubes at first with 1 mL of our samples from the 2 flasks, centrifuge it for 1 minute, and carefully pouring out the excess culture and avoid dropping the pellet (We do this for 3 times which is the same equivalent to 3 mL in total in each tubes). We would put 500 ul of multibuffer into each tubes and mix. Then, we incubate it for 5 minutes at room temperature them spin. We would repeat this another time. Finally, we are off to the kit. 

Results: 

Figure 1. 1% Gel Results


Our extraction was too low and would not show up in the gel. 









Discussion: 

    Unfortunately, we have to redo the isolation because our yield of plasmid is too low. The sample was not bright enough in the gel. Thus, instead of incubating sample and lysozyme for 30 minutes, we will now incubate for 45 minutes at 37 C. 

  




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