Revision as of 08:24, 3 October 2013 by Ggirelli (Talk | contribs)

{ "date" : "2013-07-05", "author" : "emil", "title" : " Purification of the second run of Inocula of ligation and re-ligation of the digested products ", "content" : " I added 1µl of DPN1 to the insert and 1µl of SAP to the backbones.Then I have purified the inocula following the miniprep protocol.Afterwards I quantified them toghether with the results of the purification of the digestion products(performed following the purification protocol ) with the following results:

1:1 024+0840 E482.6ng/µl
1:1 024+0840 E700ng/µl
1:1 024+0840373.1ng/µl
1:3 024+0840423.5ng/µl
1:3 024+0840327.8ng/µl
1:1 026+0840293.1ng/µl
1:1 026+08401243ng/µl
1:3 026+0840766.4ng/µl
1:3 026+0840369.1ng/µl
Then I digested 800 ng of each sample with EcoR1 HF and Pst1 following the digestion protocol with the following results:
Gel order
Ladder 1kb Fermentas1
BBa_K823024+BBa_E0840 A3
BBa_K823024+BBa_E0840 B5
BBa_K823024+BBa_E0840 C6
BBa_K823024+BBa_E0840 D7
BBa_K823024+BBa_E0840 E8
BBa_K823026+BBa_E0840 F9
BBa_K823026+BBa_E0840 G10
BBa_K823026+BBa_E0840 H11
BBa_K823026+BBa_E0840 I12
As we can see no one of the wells shows the right insert in the lower band(there is always the previous insert the RFP + Lac promoter ~ 1200 bp).At the same time I did the ligation of K823024+E0840 and K823026+E0840 previously digested following the ligation protocol .
N.B. Remember always to spin the ligation buffer before using
After that I plated the product of the ligation as done before(1:1,1:3,ctrl for each sample) on Ampicillin added LB.", "tags" : "K832024-K823026-E0840" }