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Team:Hawaii/Test Competent E. Coli
From 2008.igem.org
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# Thaw plasmid on ice | # Thaw plasmid on ice | ||
# Add 1uL plasmid pBluescript | # Add 1uL plasmid pBluescript | ||
| - | # Incubate plasmid+competent cells on ice for 30 minutes | + | # Incubate plasmid + competent cells on ice for 30 minutes |
# Heat shock 60 sec at 42C | # Heat shock 60 sec at 42C | ||
# Add 250 μl SOC | # Add 250 μl SOC | ||
# Incubate at 37 C for 1 hour in 2 ml eppendorf tubes, with shaking (150 rpm) | # Incubate at 37 C for 1 hour in 2 ml eppendorf tubes, with shaking (150 rpm) | ||
| - | # Plate 20 uL dilutions 1/1, 1/10, 1/100 to test competency | + | # Plate 20 uL dilutions 1/1, 1/10, 1/100 on LB+amp to test competency |
=== Results === | === Results === | ||
Revision as of 07:19, 4 June 2008
Contents |
Test Competent E. coli DH5a
- Test our competent cells against Callahan competent cells
Methods
Materials
- Transported 50uL of competent cells from Callahan Lab on ice
- 100uL batch 1 competent cells made at OD600 ~0.6 (split into two 50 uL aliquots 4-18a,4-18b)
- 100uL batch 2 competent cells made at OD600 ~0.3 (split into two 50 uL aliquots 2-17a,2-17b)
Procedure
- Thaw plasmid on ice
- Add 1uL plasmid pBluescript
- Incubate plasmid + competent cells on ice for 30 minutes
- Heat shock 60 sec at 42C
- Add 250 μl SOC
- Incubate at 37 C for 1 hour in 2 ml eppendorf tubes, with shaking (150 rpm)
- Plate 20 uL dilutions 1/1, 1/10, 1/100 on LB+amp to test competency
Results
Discussion
Next time, remember to aliquot out and transform only 50 ul of the competent cells.
[http://manoa.hawaii.edu/ 
][http://manoa.hawaii.edu/ovcrge/ 
][http://www.ctahr.hawaii.edu 
]
