User:Jorge Arturo Zepeda/Notebook/iGem LCG-UNAM team 2010/2010/06/17

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Thursday 17th June 2010

I performed a hot start PCR to the Green Fluorescent Protein with LVA tag for rapid degradation (BBa_K145015) to add an RBS (BBa_B0034) and 7 bases before the initial ATG, for this I used rtth polymerase.

  • 1º Reaction

Reactive.............................1x.............4.x
H2O.................................11.5ul.........46ul
Buffer 3.3x........................5ul.............20ul
Mg(OAc)2..........................3ul.............12ul
dNTP’s.............................2.5ul...........10ul
Oligo up-suffix................2.5ul...........10ul
Oligo down-preffx...........2.5ul...........10ul
DNA...................................2ul..............8ul
Total.................................30ul...........120ul

  • 2º Reaction

Reactive...............1x (ul).........4x (ul)
H2O.....................10.5............42
Buffer 3.3x............9................36
Rtth.....................0.5...............2
Total....................20...............80
The PCR came out well.