Talk:IGEM:MIT/2006/Communications: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
Line 19: | Line 19: | ||
*Funding sources for each person | *Funding sources for each person | ||
*Protocol for transformation | *Protocol for transformation | ||
See below | |||
#Transformation - André [http://openwetware.org/wiki/Transforming_chemically_competent_cells_%28Inoue%29 Transform competent cells] | |||
#*Thaw ~50 μl cells (whichever you like) on ice. Do not use glass tubes, which adsorb DNA. | |||
#*Add 200 uL DNA to cells | |||
#*Incubate on ice for 30 minutes | |||
#*Incubate cells for 50-60 seconds at 42°C. | |||
#*Incubate cells on ice for 2 min. | |||
#*Add 200-300μL of room temperature [[SOC]] (not critical) | |||
#*Incubate for 1 hour at 37°C on shaker. | |||
#*Spread 200 μl onto plates made with appropriate antibiotic. | |||
#*Grow overnight at 37°C. | |||
*Describing figure 6 | *Describing figure 6 | ||
*Update <bbpart>BBa_Q04401</bbpart> | *Update <bbpart>BBa_Q04401</bbpart> |
Revision as of 12:48, 11 February 2008
Questions for Stephen
- Did you use the Knight or Endy site directed mutagenesis protocol? (http://openwetware.org/wiki/IGEM:MIT/2006/Notebook/2006-8-10#Mutagenesis says Knight but you told me Endy)
- A: We used the Knight protocol.
- Then are you sure that you phosphorylated the primers? The Knight protocol doesn't call for this step.
- A: We used the Knight protocol.
- I'm sure we did.
- Do we have the regulated wintergreen GC data in a form that we could generate plots similar to the banana timecourse?
- Yes, we could. It would take some digging through old files and sorting out through each protocol we used on each day. I don't know if it's necessary to show though.
- What did you use to shake the cultures? i.e. what incubator or shaker?
220 RPM- the shaker in 37 C room
110 RPM- the shaker by Samantha's/2007 iGEM team's bench
General outstanding issues
- Funding sources for each person
- Protocol for transformation
See below
- Transformation - André Transform competent cells
- Thaw ~50 μl cells (whichever you like) on ice. Do not use glass tubes, which adsorb DNA.
- Add 200 uL DNA to cells
- Incubate on ice for 30 minutes
- Incubate cells for 50-60 seconds at 42°C.
- Incubate cells on ice for 2 min.
- Add 200-300μL of room temperature SOC (not critical)
- Incubate for 1 hour at 37°C on shaker.
- Spread 200 μl onto plates made with appropriate antibiotic.
- Grow overnight at 37°C.
- Describing figure 6
- Update <bbpart>BBa_Q04401</bbpart>