IGEM:Harvard/2006/Cyanobacteria/Notebook/2006-10-26: Difference between revisions
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=== Lanes === | === Lanes === | ||
# SeeBlue Plus2 | # SeeBlue Plus2 marker | ||
# J36010 (Promoter + RBS + KaiC) @ 0.55 OD | # J36010 (Promoter + RBS + KaiC) @ 0.55 OD | ||
# J36021 (Promoter + RBS + KaiB + Promoter + RBS + KaiC) @ 0.55 OD | # J36021 (Promoter + RBS + KaiB + Promoter + RBS + KaiC) @ 0.55 OD | ||
| Line 14: | Line 14: | ||
# J36022 @ 0.18 OD | # J36022 @ 0.18 OD | ||
# GFP dev (positive control) @ 0.4 OD | # GFP dev (positive control) @ 0.4 OD | ||
# SeeBlue Plus2 | # SeeBlue Plus2 marker | ||
=== Sample preparation === | === Sample preparation === | ||
Revision as of 08:53, 26 October 2006
Western blot
DUN DUN DUN
Lanes
- SeeBlue Plus2 marker
- J36010 (Promoter + RBS + KaiC) @ 0.55 OD
- J36021 (Promoter + RBS + KaiB + Promoter + RBS + KaiC) @ 0.55 OD
- J36022 (Promoter + RBS + KaiC + Promoter + RBS + KaiC) @ 0.55 OD
- J36010 @ 0.4 OD
- J36021 @ 0.4 OD
- J36022 @ 0.4 OD
- J36010 @ 0.18 OD
- J36021 @ 0.18 OD
- J36022 @ 0.18 OD
- GFP dev (positive control) @ 0.4 OD
- SeeBlue Plus2 marker
Sample preparation
At Alain's recommendation, I prepared the sample differently from the procedure in the Endy protocol.
- Spin down 1 mL of culture, freeze pellet
- Resuspend pellet in 100 uL PBS. You can freeze this for future Western blots if you wish.
- Make a 20 uL mixture containing the following:
- 5 uL sample (suspended in PBS)
- 11 uL H2O
- 4 uL 5x sample buffer
- Boil for 2 minutes
- Spin down briefly to get condensation off the top of the tube
- Insert into lanes
