Analyzing a Ligation before Transformation
I am afraid if I fail again! Oh, no! It is too bad. I did it until I cut the band from the agarose gel..but I did not do DNA isolation perfectly. Last night before, I was very tired and sleepy so that I did not see the undissolved gel! That was terrible thing I could do. I cut the gel and dissolved it with sodium iodide, incubated 37C with vigorous vortexing, even until 10 minutes. I was surprised yesterday morning when I took the tube from the cool room and then spinned it, yeah, I saw the undissolved gel! I did wanted to throw away the tube and cry..but still, I kept those tubes and added more NaI, incubated again at 4C for several hours and then continued to isolate the DNA.
After I measured the DNA concentration by spot method, I was pretty sure of the pcDNA fragment, it remained there. But I the PGV-B fragment was none, zero concentration I guessed. Just liked the last experiment I did, I continued to ligate the fragments…and did transformation. This morning I picked 2 clones up and transferred into Amp-LB-liquid. I did not have any idea except to wait until tomorrow, after I do miniprep DNA isolation and check the DNA, until I found a good idea from the internet. Yeah, it is a good idea to check the ligation before doing transformation. We also can observe the result more quickly. We have to sacrifice our sample, and of course, agarose gel (1 g of agarose is 11 Yen), but still it is a good idea.
Now, I am just waiting how it works and think how to prepare the next experiment. I can not keep on focus because I have to prepare for my progress report too.
by jamiejlg on Thu May 10, 2007 7:20 pm