Cloning of the mgtc promoter into the pSB1A3 plasmid

Kamil

Tasks:

• PCR product digest

Methods:

• The digest mix was prepared as follows:

  40μl plasmid isolation
  5μl Buffer O (Fermentas)
  1μl EcoRI enzyme
  1μl PstI enzyme
  3μl H2O

• The digest was carried out in 37°C overnight and inactivated for 15 min. in 80°C.
• The product was later purified using the Montage PCR Centrifugal Filter Device.

Cloning of the cro-box into the pSB1A3 plasmid

Kamil

Tasks:

• Creation of the cro-box double stranded DNA
• Cro-box digest

Methods:

• The mixture of equal volumes (10μl) of both ssDNA was heated to 94°C for 15min. and left to cool.
• The digest mix was prepared as follows:

  20μl plasmid isolation
  3μl Tango Buffer (Fermentas)
  1μl XbaI enzyme
  1μl PstI enzyme
  5μl H2O

• The digest was carried out in 37°C overnight and inactivated for 15 min. in 80°C.
• The product was later purified with dialysis.