E. Coli Transformation

Materials

You will need:

• DH5α competent cells
• polypropylene tubes
• ice
• ligation product
• water bath
• microtiter pipette & tips
• luria broth
• nutrient agar plates
• Sterilized glass beads

Procedure

• Thaw DH5α cells on ice for 10 minutes.
• Transfer 50μL of DH5α cells to chill, sterile polypropylene tubes.
• Transfer 1μL of ligation product into tubes.
• Incubate on ice for 30 minutes.
• Gently transfer tubes to 42°C water bath for 90s (for heat shock)
• Transfer tubes back on ice for 2 minutes.
• Add 800μL of luria broth into each tube.
• Incubate at 37°C for 1 hour in a shaker.
• Plate 200μL of each sample on nutrient agar plates. (Be sure to choose the plates with the antibiotic that your E. Coli is resistant to.)
• Spread the sample evenly by plating with sterilized glass beads. Empty the plate of glass beads afterwards.
• Incubate overnight at 37°C.

Purpose

• Notes by David Larsen

• Materials
• Procedure
• Purpose
• References

We would also like to thank and acknowledge:
Our Advisors
Marc Facciotti
Ilias Tagkopoulos
Technical Guidance
David Larsen
Andrew Yao
Visiting iGEMer
Jia Li of Zhejiang University (TEAM ZJU-China)
cI Promoter Screen
Drew Endy - Stanford
Thomas Schneider - NIH
Want to sponsor us? Send an email to mtfacciotti@ucdavis.edu to discuss various ways you can help! :)