User:Daniela A. Garcia S./Notebook/Modeling UNAM-Genomics Mexico/2010/08/02

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=Wet Lab: OmpC/F promoter=

OmpC obtaining
The OmpC promoter was obtained from the 2010 Plate [Plate 1_ Well K16]. The part is BBa_R0082 and is inserted in pSB1A2 [A=Ampicillin]


 * 15μl HPLC were added

OmpF PCR
PCR1.

Primer forward: Suffix Primere reverse: Bluepromoter-Preffix Template: pSB3K3

PCR2.

Primer forward: Suffix Primere reverse: OmpFpart1-Preffix Template: pSB3K3

PCR3.

Primer forward: Suffix Primere reverse: Bluepromoter-Preffix Template: pSB1T3

PCR4.

Primer forward: Suffix Primere reverse: OmpFpart1-Preffix Template: pSB1T3


 * General PCR mixture:
 * 4 microL Template(dilution 1:4)
 * 3 microL Primer forward (5mM)
 * 3 microL Primere reverse (5mM)
 * 6 microL HPLC
 * 6 microL 3.3X XL Buffer
 * 3 microL Mg solution
 * 5 microL dNTP's (0.4 mM)


 * Enzyme(RTTH) mixture for PCR Reaction:
 * 10.5 microL HPLC
 * 9 microL    3.3X XL Buffer
 * 0.5 microL  RTTH


 * Program [35 PCR cycles]
 * 94ºC 5min
 * 94ºC 45seg
 * 60ºC-55ºC 45seg
 * 72ºC 10min
 * 72ºC 10min
 * 4ºC 3min


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