IGEM:metu/2009/Notebook/wound dressing/2009/08/20

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20.08.2009
1. We couldn’t prepare competent cell. We missed OD. We will try again. 2. We tried to digest EGF4 with Xba1 and Pst1, GFP with Spe1 and EcoR1. Just Ecor1 digested

3. We tried to digest KGF1 with Spe1, KGF2 with Pst1. They weren’t digested 4. We think because of lacking of BSA digestion didn’t happen. 5. We prepared new leader 1 µl  leader 1 µl  10X loading dye 4 µl  dH2O

6. Electrophoresis order: 1)Leader eski / leader yeni / GFP4 (24) control / EcoR1 / Spe1 / GFP (28) control / EcoR1  / Spe1   2)EGF4(6) / Pst1(1)  /Pst1(2) / Xba1 / EGF4(11) / Pst1 / Pst1(2) / Xba1 7.The following amounts were added to microtubes. Monodigestion (fast);

KGF1(AI2-1),  monodigestion: water 14.09 µl Buffer 2 µl DNA    2.91 µl Spe1   1 µl    =20 µl KGF1(AI2-3), monodigestion: water 14.13 µl Buffer 2 µl DNA    2.87 µl Spe1   1 µl    =20 µl KGF2(rbs1), monodigestion: water 13.41µl Buffer 2 µl DNA    3.59 µl Pst1   1 µl     =20 µl KGF2(rbs2), monodigestion: water 13.99 µl Buffer 2 µl DNA    3.01 µl Pst1   1 µl     =20 µl EGF1(06.08.09), monodigestion: water 14.02 µl Buffer 2 µl DNA    2.98 µl Spe1   1 µl    =20 µl EGF1(11.08.09), monodigestion: water 14.89µl Buffer 2 µl DNA    2.11 µl Spe1   1 µl    =20 µl

EGF2(rbs1), monodigestion: water 13.41µl Buffer 2 µl DNA    3.59µl Pst1   1 µl    =20 µl

EGF2(rbs2), monodigestion: water 13.99 µl Buffer 2 µl DNA    3.01 µl Pst1   1 µl    =20 µl 8. Electrophoresis order: 1)Leader / rbs1 control / rbs1- / rbs1 / rbs2 control / rbs2- / rbs2 / leader   2)EGF1 control (06.08.09) / EGF1 cut  / EGF1 control (11.08.09) / EGF1 cut / KGF1CONTROL (AI2-1) / AI2-1 cut / AI2-3 control  / AI2-3 cut