Post on 13-Dec-2015
National Centre for Biotechnology Education
www.ncbe.reading.ac.uk
Bacterial
Transformation
Copyright © Dean Madden, 2013
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The bacteriumBacterial
chromosome
Plasmids
Ribosomes
Cell wall
Cell membranes
Cytoplasm
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GFP geneThis gene encodes green fluorescent protein, which glows in UV light
Kanamycin resistance genethe enzyme encoded by this gene stops the antibiotic kanamycin from working
Origin of replicationthis is needed so that the plasmid can be copied, using the bacterium’s DNA copying machinery
The plasmid
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Aequorea victoria
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GFP
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13Glo Fish
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Stock bacteria
These should be grown 1–2 days in advance
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Twiddle vigorously to dislodge the bacteria and
disperse them
Transformation buffer
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Leave on ice for 15 minutes to recover
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Bacterial cell with pores in cell membrane
GFPplasmid
DNA
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LB agar+ kanamycin
+ IPTG
3 drops(100 µL)
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Label the base of each plate with your initials, the
date and the type of bacteria used
Seal the plates with
tape
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BIOHAZARD
TRANSFORMED CELLS MUST BE DESTROYED
AFTER USE