Team:Paris/July 30

Analysis of yesterday DNA digestion
The digested DNA of yesterday was analysed one more time by electrophoresis on a 0.8% agarose gel (about 30 minutes at 100 W).
 * The ladder used was the 1 kb DNA ladder (New England Biolabs).
 * 5 µL of each sample with 1 µL of loading dye were loaded.'



==> Conclusion :Each of the samples was succesfully digested and purified except for the sample D108. It seems that the QIAprep columms (from the QIAGEN Minipreps kit) can be used instead of the QIAquick columms (for DNA Gel Extraction).

PCR Screening of Ligation Transformants
Use of 8 clones of Ligation transformants for screening PCR

Protocol of screening PCR

 * Mix


 * 50µl of Mix PCR by tube/clone
 * one toothpick of each clone's colony by tube
 * Program : Annealing 55°C - Time élongation 1'30" - Number cycle : 29

Conditions of electrophoresis

 * 10µl of ladder 1 kb
 * 15µl of screening PCR (gel n°1, 2, 3(9-17), 4, 5, 6, 7, 8, 9, 10, 11)
 * 10µl of screening PCR (gel n°3(1), 13, 14)
 * migration ~30min at 100W on 0,8% gel

Results
















==> Conclusion : with the PCR, we have check that the transformant bacteria contain insert. (obtain amplification at the good size).

But we don't observe results for L102(3), L102(6), L103(4), L106(1), L106(2), L106(4), L111(1)

Migration of an another gel for this sample...

Results:



==> Conclusion : We can observe a results for the samples : L102, L103, L106(4) and L111. (but not for L106(1; 2))