November 11th- November 15th

 Week 8 (Nov 11th- Nov15th)


This week, we began the discussion of using lambda DNA to experience with as a control group. Because our past processes using xbar has been unsuccessful, we have decided to use experimental DNA in order to go through a step-by-step sleuth to see which step is failing. There’s also the possibilities that our ligation is not working but digestion is, our primers are bad, and so forth. Essentially, the major question is: is it us, or our processes?


We decided that it was significant to keep our processes with lambda DNA as exact as possible in order to be able to compare the methods the most accurately. If the processes are not constant like before, there’s no point at all in doing the experiment.



Lambda key data:

  Lambda DNA: #N3011S, New England BioLabs

500 ug/ml

48,502 base pairs

Xbar site at 24508 which leaves a 23994 bp difference


The biggest issue with this is that the base pair difference is so tiny, sitting only at a 514 difference, therefore it’ll be extremely hard to tell them apart on the gel.


We decided to run our gel for around two hours so that we could get the most clear and concise answer as possible as opposed to not being able to read the slight differences between the DNA fragments.


We ended up making a 2% Gel, using 120 ml instead of 30 ml. On our gel, we are going to put our ladder, our lambda-digested DNA, and plain lambda dna so we can see if it digested.


For our digest of lambda, we used 2 ul buffer, 2ul dna, 1 ul enzyme, and 15 ul of water.


After overnight digestion, repeating the process that we had done before, we ran it on our gel and by the end of our two hours decided that because the base pairs were both at the very top and with no difference, digestion using xbar did not occur. We did decide that we may need to redo this gel to assure 100% what has happened since there is worry that we were not able to run the gel long enough and at a high enough amp.

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