Long Term Goals
miRNA Profile
- Read literature
- Choose profile
Building the Cell Detector Circuit
- Build Choose profile
- Verify sensors
- Create mathematical model
- Make test bed
- Controlled miRNA expression
- Constructed miRNA sensors
Mini experiments
- Single miRNA detector circuit
Experiments
- L7ae repression
- TAL14 repression
- TAL21 repression
- Single input sensors
- Multiple input sensors
Alzheimer's Model
- Agree on a team wide, full Alzheimer's model
Today's Agenda
How To Please Ron/Brain
- Create a clear workflow
- Update "status" of plasmids on parts list
- How do we string multiple low sensors together?
- Keep updating the parts tree (include experiments)
miRNA Sensors (Theory)
- Determine which pairs of target sites we want
- Find sequences (hopefully under 200 bp)
- Send ultramer order to Brian
- Learn how to acquire MATLAB from Brian
- Find natural concentrations of relevant miRNAs.
miRNA Sensors (Lab Work)
- Tranform LRs
- MAV1212-Hef1a-mKate2
- MAV1212-Hef1a-L7ae
- Hef1a on Kan
- Pick colonies from transformations
- MAV1212-Hef1a-eBFP2
- Golden Gate
- Gather miRNA target sites from Jeremy
- Make all 6 single low sensors
NOTES
MAV1212 didn't grow (ccdB resistance is over expressed)
Hef1a was plated on Amp plates...but it has Kan resistance
- Midi-prep hEF1a:rtTA and hEF1a: Gal4VP16 2A rtTA3
- Mini-prep MAV1212 hEF1A Tal14
- Digest and Map MAV1212 hEF1A Tal14
- Transform third LR trial if not verified
- Pick/Streak hEF1a: Gal4VP16 2A rtTA3
NOTES:
hEF1a: Gal4VP16 2A rtTA3 culture did not grow so Verify MAV1212-Hef1a-eBFP2 via sequencing AmpR/eBFP2 region