User:Wilfred J. Poppinga/Notebook/Wilfreds Project/2009/08/25

Combining MBP-ArsR with inducible promotors

• Cut plasmids containing promotor with SpeI and PstI
  • 1 μL 10x fast digest buffer
  • 0.5 μL SpeI
  • 0.5 μL PstI
  • 8 μL Vector
• Incubate 1 h @ 37 °C
• Purify cut plasmid using PCR clean up kit

• Cut plasmids containing MBP-ArsR with XbaI and PstI
  • 1 μL 10x fast digest buffer
  • 0.5 μL XbaI
  • 0.5 μL PstI
  • 8 μL Vector
• Incubate 1 h @ 37 °C
• Put restriction on 1% Agarose (TBE) gel
  • Cut band of ~1500 bp out
  • Purify using gel purification kit
    • End volume 30 μL

• Ligation
  • μL ligation buffer
  • 1 μL ligase
  • μL vector
  • μL insert
  • Incubate

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