Transformation of Bacteria: Difference between revisions
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updated transformation protocol |
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==Materials== | ==Materials== | ||
*Competent Cells | *Competent Cells | ||
*SOC Buffer or LB Media | |||
*SOC Buffer | |||
*DNA of interest (1 uL for plasmid, 5 uL for cloning/mutagenesis) | *DNA of interest (1 uL for plasmid, 5 uL for cloning/mutagenesis) | ||
*Plates (Amp or Kan; in cold room) | *Plates (Amp or Kan; in cold room, see [[ Making LB Agar Plates ]]) | ||
==Protocol== | ==Protocol== | ||
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#Heat shock at 42C for 45s | #Heat shock at 42C for 45s | ||
#Place back on ice | #Place back on ice | ||
#Add 450 uL of SOC Buffer or LB | #Add 450 uL of SOC Buffer or LB. | ||
#Incubate at 37C for 1h with occasional mixing | #Incubate at 37C for 1h with occasional mixing | ||
#Plate 30-50 uL for amplification, or all for cloning/mutagenesis | #Plate 30-50 uL for amplification, or all for cloning/mutagenesis | ||
*When plating be sure to label plates and use extreme caution when sterilizing the spreader with EtOH and fire. | *When plating be sure to label plates and use extreme caution when sterilizing the spreader with EtOH and fire. | ||