"

From 2013.igem.org

{ "date" : "2013-06-18", "author" : "gabriele-emil", "title" : "SAMsynthetase: episode 2
Attack of the Insert", "content" : "This morning we added 1µl of SAP to the pSB1C3 overnight digestion and 1µl of DpN1 to the SAMsynthetase overnight digestion, then both were incubated at 37°C for 1.5 hours.

During these 1.5 hours, we performed the miniprep (protocol) and quantification of the circular pSB1C3 inocula of yesterday.

After the incubation, we purified the digestion mixes using the Wizard® SV Gel and PCR Clean-Up System and then quantified.
Then, we performed the ligation of pSB1C3 and SAMsynthetase exploiting the usual protocol.We incubated the ligation mixes for 2 hours at room temperature.

Also, we extracted R0010 promoter (Plac) from the registry (2013 distribution kit, plate 3, well 3H). Unfortunately, we also extracted a part from the same plate and well of the 2012 distribution kit (BBa_K115032) because we mistook the kits.

Finally, we transformed NEB10β competent cells with the usual protocol, we used the following quantities of DNA: 10µl of each ligation product except for the 1:4 ligation, of which we used 15µl, and 1µl of the extracted R0010. Then, we plated on CM plates.", "tags" : "SAMsynthetase" }