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Team:NTU-Singapore/Notebook
From 2009.igem.org
Experiment Timeline
June
18th June
- Preparation of LB medium
- Amplification of parts from registry (pTet, TetR, HO1, Low copy plasmid)
19th June
- Antibiotic stock preparation
- Inoculation
20th June
- Our first Plasmid Extraction (PE)!
22nd June
- First Double Digestion (DD) and Gel Check!
23rd June
- PE & DD of the registry constructs….
24th June
- Our direct synthesized parts finally arrived. We now have CHE (cholesterol esterase), IFP(Infrared protein 1.4) and pNO (NO sensing promoter).
26th June
- PE , DD and Gel check for the direct synthesized gene sequence.
27th June
- Gel Check for CHE, IFP and pNO
July
1st July
- PE for plaqIQ, J23101, J23119, B0034(rbs), B0015(Term-T for short), B0034-GFP-B0015
2nd July
- PE for pNO, IFP, CHE
- DD of pNO, J23119, J23101, rbs-GFP-T
- Our first ligation!
3rd July
- DD of CHE, IFP and B0015, plaqIQ and plux.
- Ligation.
4th July
- PE: pNO-GFP; J23101-GFP; J23119-GFP; HO1
- DD: HO1
7th July
- PE: CHE, IFP, placIQ
- DD:CHE-T, IFP-T. HO1, Term
- Ligation
8th July
- DD: pNO-GFP; J23101-GFP; J23119-GFP
9th July
- PE
- DD: ptet. r-CHE-T, r-HO1, r-IFP-T
- Ligation: r-HO1 +r-IFP-T
10th July
- PE
- DD: ptet; r-CHE-T, Term, tetR
11th July
- PE
- DD: r-CHE-T; r-HO1-r-IFP-T;CHE-T; r-CHE-T
13th July
- DD: ptet; r-CHE-T; r-HO1-r-IFP-T; TetR; Term
- Ligation: ptet + r-CHE-T; ptet + r-HO1-r-IFP-T; TetR + Term
15th July
- PE
- DD: pTet; rbs; IFP; CHE; TetR-T
16th July
- DD: pTet; r-CHE-T; Inv; r-HO1-r-IFP-T; Term; TetR
- Ligation
18th July
- PE
- DD and Single Digestion (SD) :r-HO1-r-IFP-T (because they have the same size)
20th July
- DD:pTet; J23119; r-HO1-r-IFP-T; TetR; Term; placIQ; inv
- Ligation: pTet +r-HO1-r-IFP-T; J23119 + r-HO1-r-IFP-T; placIQ +Inv; TetR +Term
- PE
23rd July
- PE
- DD:J23119; Inv; r-HO1; r-GFP-T; pTet; placIQ-Inv; TetR-T
24th July
- PE
- DD: J23119; r-CHE-T; pTet; inv;r-HO1-r-IFP-T; ptet-IFP
- Ligation: J23119+r-CHE-T; ptet+r-CHE-T; ptet+r-HO1-r-IFP-T; J23119+Inv
28th July
- PE
- DD: r-HO1; r-GFP-T; J23119;r-CHE-T;r-HO1-r-IFP-t; ptet; r-CHE-T
- Ligation: r-HO1 +r-GFP-T; J23119 +r-CHE-T; J23119 + r-HO1-r-IFP-T; pTet +r-CHE-T; pTet +r-GFP-T
30th July
- PE
- DD : J23119-r-CHE-T; ptet-r-CHE-T; J23119-r-HO1-r-IFP-T; ptet-r-HO1-r-IFP-T; r-HO1-r-GFP-T; J23119; pTet; TetR; rbs
- Ligation: r-HO1 +r-GFP-T; J23119 +r-CHE-T; J23119+r-HO1-IFP-T; ptet+r-CHE-T; ptet+r-GFP-T
August
4th August
- DD :rbs; NorR
5th August
- PE
- DD
- Ligation
12th August
- DD
- Ligation
14th August
- DD
- Ligation
17th August
- DD
- Our first SDS-PAGE—J23119-r-CHE-T; pTet-r-CHE-T
18th August
- DD
- Ligation
21st August
- DD
- Characterization of J23119
24th August
- PCR verification –HO1, pTet constructs
25th August
- pTet characterization
26th August
- DD
27th August
- DD
- Ligation
28th August
- PCR verification – CHE constructs
- DD
- Ligation
29th August
- DD
- Ligation
- PCR verification –IFP constructs
31st August
- PCR verification –HO1, CHE constructs
- DD
- Ligation
September
1st September
- PCR verification –IFP, NorR, pNO
- DD
- Ligation
2nd September
- DD
- Ligation
3rd September
- DD
- Ligation
4th September
- DD
- Ligation
6th September
- PCR --J23119-r-NorR-T
7th September
- DD,
- Ligation
8th September
- PCR verification –IFP, HO1 constructs
9th September
- PCR verification –HO1 constructs
10th September
- DD
- Ligation
11th September
- PCR verification –NorR, IFP, HO1 constructs
- Ligation
12thS eptember
- PCR verification – NorR, IFP, HO1 constructs
14thSeptember
- PCR verification – IFP, HO1 constructs
15th September
- DD
- Ligation
- SDS-Page
16th September
- pNO characterization
17th September
- Ligation
18th September
- DD
- Ligation
19th September
- DD
20th September
- Ligation
22nd September
- PCR verification –IFP, HO1
- pNO characterization
23rd September
- PCR verification --CHE
24th September
- PCR verification –CHE, HO1
- DD
- Ligation
- Optimization of CHE characterization experiment
26th September
- DD
- Ligation
