Welcome to the Creation of the Cloning Vector Page!

Background:

external image 500px-L4440.tif.png
Procedure:
1. 2 cultures needed to make mini-preps:
  • LB tubes
  • AMP (1,000 x) 1 uL/mL
  • ~500 uL L4440 (in DH5alpha) per culture

2. Place the cultures in a shaking water bath for 24 hours.

3. Conduct 4 Zippy mini-preps to separate the L4440 plasmid from the E. coli DH5alpha strain host cells.

4. Digest the L4440 vector using the following protocols.
  • 1 ul of each enzyme (HindIII, SacII)
  • 10 ul L4440 vector
  • 3 ul 10x buffer
  • 15 ul dd water

5. Put in a 37 degrees Celsius water bath.