Week 1

Friday

Today we started with a briefing from Mustafa where the main action points of the day were discussed. We had been planning the project of this year.We determined our parts that will be necessary:

• K124014 holin
• K559010 halorhodopsin+terminator(X2)
• K112806 endolysine/T4
• K112808 endolysine/holin/CMV/antiholin

These 4 parts was diluted and tagged. Then we coded these parts:

• C1:J23100 cons-promoter
• C2:CMV promoter
• P1:CL promoter
• H1:Holorodopsin
• P2:IPTG
• L1:endolsin
• L2:holin
• l3:eth+antiholin


We prepared 35 plates and 200 ml LB broth for next experiments.

• Transformation (L1,L2,L3) 

• C1,C2 streak
• Coloniese are incubated in liquid culture.(C1)

Saturday

• Coloniese are incubated liquid culture.( 1,5 ml AMP ) for :

C1(x2),C2,N1,L2,L3.

• We made this process 2 times because of high mistake risk for first times

Sunday

• Single colony isolation has made for C1-x,C1-y,C2,L1,L2 and L3.
• C1-x and C1-y are digested with EcoR1 and Pst1.
• We prepared liquid culture for C2,L1,L3 again.Because there was not enough DNA for digestion.

Monday

• Single colony isolation has been made for L1,L2 and L3.
• Digestion has been made with EcoR1 and Pst1 for L1,L3 and C2.
• electrophoresis was performed for C1-x,C2,L1,L2 and L3.
• After electrophoresis,results of L1,L3 was true but the others was wrong.

Tuesday

• Digestion has been made with Xba1 and Pst1 for L1 and L3.
• 34 mg/ml chloramphenicol included 49 tubes was prepared.
• Chloramphenicol included 17 plates was prepared.
• IPTG induceble promoter was coded as P2.
• Transformation has been made for H1,L1P2 and L3P2
• We prepared liquid culture for C1,C2,L2

Wednesday

• Single colony isolation has been made for C1,C2,L2
• We prepared liquid culture for C2 again.Because there was not enough DNA for digestion.
• C1 and L2 are digested.
• electrophoresis was performed for C1 and L2.
• The result of C1 was true but the result of L2 was wrong so a new digestion will been performed and after that electrophoresis will been performed for new L2 and old L2.
• We prepared liquid culture for P2L1,P2L3,C2,H1 and L2.

Thursday

• We made isolation for P2L1,P2L3,H1,C2,L2 and then digestion has been made with EcoR1 and Pst1 for all of these parts.After that electrophoresis was performed for these parts.
• The results of P2L1 and P2L3 might be true , we are not sure about this so we need some expert assistance and we need to sequence these part for being sure. Unfortunately the result of L2 was wrong again.
• We prepared a new liquid culture for L2.