Team:Groningen/Notebook/5 August 2009

From 2009.igem.org

(Difference between revisions)
m (GVP Cluster)
(GVP Cluster)
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:→ {{todo}} isolate plasmids from overnight precultures
:→ {{todo}} isolate plasmids from overnight precultures
:→ {{done}} perform a ligation between cut vector and GVP fragment
:→ {{done}} perform a ligation between cut vector and GVP fragment
-
:→ {{todo}} transform E.coli TOP10 competent cells with ligation product
+
:→ {{done}} transform E.coli TOP10 competent cells with ligation product
'''Ligation'''
'''Ligation'''

Revision as of 11:33, 5 August 2009

Igemhomelogo.png

It's my birthday!!! (Michael)

So cake it is....

Wet

GVP Cluster

TODO buy a nice cake and eat it ;)
TODO isolate plasmids from overnight precultures
DONE perform a ligation between cut vector and GVP fragment
DONE transform E.coli TOP10 competent cells with ligation product

Ligation

  • 2 uL Ligase buffer
  • 1 ul T4 Ligase
  • 8 uL plasmid pSB1AC3 digested with PstI and SpeI
  • 4 uL insert GVP restricted with XbaI and PstI

Incubate:

  • 25°C 30min.
  • kept on ice for 10min.

Tranformation

  • add 10uL of the pSB1AC3-GVP ligation product to 50uL competent E.coli TOP10 cells.

Incubate:

  • 30 min @ ice
  • 50 sec 42°C
  • 2 min @ ice
  • add 800uL LB-medium
  • incubate for 1 h at 37°C
  • plate on LB-amp50 plates

Dry

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