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Created by Unknown User (lylaatta@mit.edu), last modified by Unknown User (clrichar@mit.edu) on May 29, 2015 22:03
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Experimental Progress:
Linker Optimization
Useful Links:
Linker optimization
Task | Claimed by | Date Completed | Other Notes |
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CD79A/B, Gal4VP16, RFP PCR try3 | CR | Apr 10 | |
Gel verification of try3 PCR product | LA | Apr 10 | RFP and Gal4 worked, CD79s didn't - we probably need to redesign primers |
Redesign primers | LA + CR | Apr 11 | CD79A F and R and CD79B F primers were redesigned and ordered |
AarI golden gate of CD79A construct | LA | Apr 30 | |
Take out CD79A golden gate plates | LA | May 2 | |
Make LB | CR | May 4 | |
CD79A golden gate liquid cultures | LA | May 4 | Cultures in at 6pm - need to be taken out by 10/11 am May 5 in 215 shaking incubator |
Take out CD79A golden gate liquid cultures | LA | May 5 | In 4C |
CD79A golden gate minipreps | LA | May 6 | In yellow rack in 4C - need to be nanodropped |
CD79A golden gate restriction verification | CR | May 6 | Some promising constructs |
Send promising CD79A golden gates for sequencing | | | |
- CD79A linker golden gates
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Redesign CD79B reverse PCR primer | CR | May 24 | |
Variable region construction
Task | Claimed by | Date Completed | Other Notes |
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Order primers for PCRing constant region | | | primers on geneious |
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Christian is Confused so he is using this to do list
Day | List |
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Monday |
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Tuesday | Design CD79B Sequencing PrimersLook into tTEV and IGD optionsApply for SynBerc and BDACPlan Lyn and Syk ExperimentPlan mAB cloning
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Wednesday | Establish HEK293 Cell LineEstablish Cryostocks of Gmab H, Gmab L, Syk-TEV, Lyn-TEV, CD79A, CD79B, CD79A-G4, CD79B-G4, hEF1a: CD79A - pTet: RFP - Gal4VP16Transfect Lyn/TEV experiment
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Thursday | Sequence hEF1a: CD79A - pTet: RFP - Gal4VP16PCR CD79B with third gen primersDigest & Gel CD79B &hEF1a: CD79A - pTet: RFP - Gal4VP16GG hEF1a:CD79B-pTet:RFP-Gal4VP16GG H:CDA-RFP-G4 with linkers
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Friday |
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Saturday | Pick Colonies hEF1a:CD79B-pTet:RFP-Gal4VP16Pick colonies H:CDA-RFP-G4 w/ linkersSplit HEK293
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Sunday | Miniprep hEF1a:CD79B-pTet:RFP-Gal4VP16Miniprep H:CDA-RFP-G4 W/ LinkersDigest & Gel hEF1a:CD79B-pTet:RFP-Gal4VP16Digest & Gel H:CDA-RFP-G4 W/ Linkers
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Day | List |
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Monday |
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Tuesday |
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Wednesday |
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Thursday |
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Friday | Inoculate 5mL CulturesPCR CD79BGel CD79BSequence H:CDA-RFP-G4
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Saturday | Inoculate MidiprepsPrepare CryostocksPick Colonies => Midipreps & 5mL for Glycerol StocksPour New GelsGG H:CDB-RFP-G4Check for Sequencing for H:CDA-RFP-G4Check HEK Cells
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Sunday |
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Higher Level To Do's:
- Finish Discussing Figures for Experiment list
- Decide on other antigen-variable regions we want to test
- Syk-Lyn experiment design
- Receptor dilution experiment design
- Other experimental planning