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| Date | Summary |
|---|
| 07/01 | - hEF1a:CD79B-pTET:mRFP-Gal4VP16 CD79B mutation
- pLac:mKate miniprep, digest, gel
- TRE:syk-pTet:mRFP-(t)TEVp miniprep, digest, gel
- hEF1a:CymR, hEF1a-CuO:mKate, CMV5-CuO:mKate LR
- pENTR LacI, pENTR hEF1a-LacO transformations
- TRE:Gal4VP16, TRE:mKate, hEF1a:LacI midi culture
|
07/02 | |
Spring
| Date | Summary |
|---|
| 03/23 | Verification of linker-switchable PCRs |
| 04/01 | - rePCRs of CD79B, Gal4 and RFP to make linker switchable constructs
- Gel verification of 03/31 and 04/01 PCRs
|
| 04/10 | - rePCRs of CD79A/B, Gal4 and RFP to make linker switchable constructs
- Gel verification of PCRs
|
| 04/11 | redesign CD79A/B primers |
| 04/25 | - rePCRs of CD79A/B, to make linker switchable constructs
- Gel verification of PCRs
|
| 04/30 | - rePCR of CD79B with DMSO
- gel purification of CD79A
- PCR purification of pTet:mRFP and Gal4VP16
- AarI golden gate to make hEF1a:CD79A-pTet:mRFP-Gal4VP16
|
| 05/01 | |
| 05/04 | |
| 05/06 | |
| 05/29 | - Transformations for Glycerol Stocks
- Sent for hEF1a:CD79B-TCS-Gal4VP16 sequencing
- PCRd CD79B
- CD79B Gel
|
| 05/30 | - CD79B Gel Extraction (Failed)
- Midi Cultures Inocculated
- CD79B PCR 2
- Picked Colonies for H:Lyn, H:LT, H:CB-TCS-G4
|
| 05/31 | - CD79B PCR Gel
- CD79B Gel Extraction
- hEF1a:CD79B-pTet:RFP-Gal4VP16 Golden Gate
- h:Lyn-TEV colonies picked
|
June
| Date | Summary |
|---|
| 06/01 | - h:Lyn-TEV Colonies Picked
- h:CDB-RFP-G4 transformation
- h:CDA-RFP-G4 Sequence Analysis
- Inocculate Midipreps
- h:Lyn, h:CDB-TCS-G4 cryostocks
|
| 06/02 | - Cloning Redesign
- Pick hEF1a:CD79B-pTet:RFP-Gal4VP16 Colonies
|
| 06/03 | - Miniprep hEF1a:CD79B-pTet:RFP-Gal4VP16
- Cryostock hEF1a:CD79A-pTet:RFP-Gal4VP16
|
| 06/04 | - Glycerol Stock hEF1a:CD79A-pTet:RFP-Gal4VP16
- Midiprep 4 Frozen pellets
- Digest & Gel hEF1a:CD79B-pTet:RFP-Gal4VP16
|
| 06/08 | - hEF1a:CD79B-pTet:RFP-Gal4VP16 Digest & gel
|
| 06/09 | |
| 06/11 | - Sequencing Analysis
- Glycerol Stock H:CDA-RFP-G4
|
| 06/16 | - designed SDM primers
- LR TRE:mKate, hEF1a:LacI, pLac:mKate
- experimental design + deadlines
|
| 06/17 | - annealed linker halves
- CD79A linker golden gates
|
| 06/18 | - transformed CD79A linker golden gates
- transformed LRs (TRE:mKate, pLac:mKate, hEF1a:LacI)
- LRed TREt:VP16Gal4
|
| 06/19 | - sdm primers are here
- killed LR TREt:VP16Gal4
- Transformed LR TREt:VP16Gal4
- Re-transformed CA-9T6-G4
- picked CD79A linker colonies
|
| 06/21 | - picked CA-9T6-G4 colonies
|
| 06/22 | - mini CD79A linkers
- mini TRE:mKate, pLac:mKate, hEF1a:LacI
- digest
- run CD79B SDM gel
|
| 06/23 | - digests and gels for TRE:mKate, pLac:mKate, hEF1a:LacI
- kill TRE:Gal4VP16 LR
- transform TRE:Gal4VP16 LR
- run SDM on gel -> gel extraction
- CD79B-RFP-G4 SDM product transformation
|
| 06/24 | - SDM CD79B-RFP-Gal4
- SDM DpnI digest
- transform SDM product
- transform TRE:Gal4VP16
|
| 06/25 | - redigest TRE:mKate, pLac:mKate, hEF1a:LacI
- PCR syk, tevp, ttevp
- gel verify PCRs and digested LRs
- TRE:gal4VP16 pick colonies and minipreps
- PCR purification of syk, tevp, ttevp
- reLR of TRE:mKate, pLac:mKate, hEF1a:LacI
- retransform SDM digests
|
| 06/26 | - TRE:gal4vp16 gel
- TRE:syk-mrfp-tevp golden gates
|
| 06/28 | |
| 06/29 | - cultured and miniprepped LR2 TRE:mKate, pLac:mKate, hEF1a:LacI
- miniprepped LR1 TRE:mKate, pLac:mKate, hEF1a:LacI
- digest and gel LR1 TRE:mKate, pLac:mKate, hEF1a:LacI
- picked colonies of new TRE:gal4vp16 transformation
|
| 06/30 | |