Team:Pittsburgh/Notebook
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Our weekly progress. For a list of our protocols, visit the Protocols page
Week 1: May 23 - May 27
Wet Lab
- Training begins
- Grow Top 10 competent cells.
Dry Lab
- Brainstorm genetic circuits for a thallium sensor
- Lab safety training
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Week 2: May 31 - June 3
Wet Lab
- Test efficiency of competent cells
Cell-Free Extract
- Test cell-free extract reaction with T7-GFP plasmid
Dry Lab
- Contact museums and summer programs for outreach opportunities
- Lab safety training
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Week 3: June 6 - June 12
Wet Lab
Reporter
- Transform T7 promoter, amilCP, and terminator
- Begin assembly by ligating linearized T7 promoter and amilCP
Dry Lab
- Contact museums and summer programs for outreach opportunities
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Week 4: June 13 - June 17
Wet Lab
Reporter
- Transform T7-GFP plasmid, lacZ alpha fragment, and eGFP
- Send promising T7 promoter -- amilCP ligations to be sequenced
- Perform double digest of T7 promoter and terminator from last week
- Ligate double-digested T7 promoter to new reporters (lacZ and eGFP)
Dry Lab
- TECBio, DiSCoBio, and Tissue Engineering Camp outreach opportunities set
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Week 5: June 20 - June 26
Wet Lab
Reporter
- Ligate T7 promoter -- amilCP construct to terminator
- Extract successful ligations of T7 promoter to eGFP
- Ligate T7 promoter -- eGFP construct to terminator
Cell-Free Extract
- Reaction volume can be reduced to 5 μL with 5 ng/μL of DNA
Toehold Switch
- Collins triggers activate the switches (both in plasmid form) to express LacZ
Dry Lab
- Reach out to teams to collaborate based on last year's projects
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Week 6: June 27 - July 3
Wet Lab
Reporter
- Identify successful ligations to terminator for amilCP and eGFP consturcts using a gel
- Send correct plasmids for sequencing for confirmation
- Test plasmids in cell-free extract
- amilCP does not produce color in cell-free reaction
- eGFP produces fluorescence comparable to that from the Collins T7-GFP plasmid in cell-free reaction
- Linearized plasmids containing only the promoter and insert (no terminator) do not express protein
Cell-Free Extract
- Reaction volume can be reduced to 1 μL with 5 ng/μL of DNA
Toehold Switch
- Collins plasmids express LacZ with 25 ng of switch
- DNA oligos trigger Collins switches
Dry Lab
- Work on outreach presentation for tissue engineering camp
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