Team:Heidelberg/Templates/Del week16 AG

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Contents

12-08-2013

Amplification from FS_02 to FS_11; 22.8 kb

Amplification of DelAG (12-08-13); run at 100 V, 0.8 % gel (TAE)
Reaction
Reagent DelAG
Template D.acidovorans SPH-1 colony
Primer fw 4.5 µl FS_02
Primer rev 4.5 µl FS_11
DMSO 1 µl
Phusion Ready Mix 10 µl


Conditions
Biometra TProfessional Basic
Cycles temperature [°C] Time [s]
1 98 10
12 98 1
72.0 - 67.0 (ΔT = 0.5) ↓ 0.5 5
72 7:20
18 98 1
70.0 - 65.0 (ΔT = 0.5) 5
72 7:20
1 72 10min
1 10 inf

Results:

  • none of the amplifications led to yields of DNA sufficient for Gibson Assembly, therefore the used combination of primers seems not to be sufficient for amplification of 22.0 kbp

13-08-2013

Amplification from FS_02 to FS_11_short; 22.8 kb

Reaction
Reagent DelAG
Template D.acidovorans SPH-1 colony
Primer fw 4.0 µl FS_02
Primer rev 4.0 µl FS_11_short
DMSO 2 µl
Phusion Ready Mix 10 µl


Conditions
Biometra TProfessional Basic
Cycles temperature [°C] Time [s]
1 98 10
12 98 1
72.0 - 67.0 (ΔT = 0.5) ↓ 0.5 5
72 7:20
18 98 1
70.0 - 65.0 (ΔT = 0.5) 5
72 7:20
1 72 10min
1 10 inf