Team:Korea U Seoul/Notebook
From 2010.igem.org
(→Brain storming & Work notes) |
(→Brain storming & Work notes) |
||
Line 7: | Line 7: | ||
== Brain storming & Work notes == | == Brain storming & Work notes == | ||
Click on a date to see notes on the meeting & summary of labwork done on that day. | Click on a date to see notes on the meeting & summary of labwork done on that day. | ||
- | + | | align="center" | |
|-valign="top" | |-valign="top" | ||
|align="center" width="150pt"|{{#calendar: title=Korea_U_Seoul |year=2010 | month=06}} | |align="center" width="150pt"|{{#calendar: title=Korea_U_Seoul |year=2010 | month=06}} |
Revision as of 12:05, 27 October 2010
![](https://static.igem.org/mediawiki/2010/4/49/KU.jpg)
Brain storming & Work notes
Click on a date to see notes on the meeting & summary of labwork done on that day. | align="center" |-valign="top" |align="center" width="150pt"|
June | ||||||
M | T | W | T | F | S | S |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/1_June_2010&action=edit 1] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/2_June_2010&action=edit 2] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/3_June_2010&action=edit 3] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/4_June_2010&action=edit 4] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/5_June_2010&action=edit 5] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/6_June_2010&action=edit 6] | |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/7_June_2010&action=edit 7] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/8_June_2010&action=edit 8] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/9_June_2010&action=edit 9] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/10_June_2010&action=edit 10] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/11_June_2010&action=edit 11] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/12_June_2010&action=edit 12] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/13_June_2010&action=edit 13] |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/14_June_2010&action=edit 14] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/15_June_2010&action=edit 15] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/16_June_2010&action=edit 16] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/17_June_2010&action=edit 17] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/18_June_2010&action=edit 18] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/19_June_2010&action=edit 19] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/20_June_2010&action=edit 20] |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/21_June_2010&action=edit 21] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/22_June_2010&action=edit 22] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/23_June_2010&action=edit 23] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/24_June_2010&action=edit 24] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/25_June_2010&action=edit 25] | [http://2010.igem.org/Korea_U_Seoul/26_June_2010 26] | [http://2010.igem.org/Korea_U_Seoul/27_June_2010 27] |
[http://2010.igem.org/Korea_U_Seoul/28_June_2010 28] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/29_June_2010&action=edit 29] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/30_June_2010&action=edit 30] |
|align="center" width="150pt"|
July | ||||||
M | T | W | T | F | S | S |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/1_July_2010&action=edit 1] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/2_July_2010&action=edit 2] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/3_July_2010&action=edit 3] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/4_July_2010&action=edit 4] | |||
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/5_July_2010&action=edit 5] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/6_July_2010&action=edit 6] | [http://2010.igem.org/Korea_U_Seoul/7_July_2010 7] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/8_July_2010&action=edit 8] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/9_July_2010&action=edit 9] | [http://2010.igem.org/Korea_U_Seoul/10_July_2010 10] | [http://2010.igem.org/Korea_U_Seoul/11_July_2010 11] |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/12_July_2010&action=edit 12] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/13_July_2010&action=edit 13] | [http://2010.igem.org/Korea_U_Seoul/14_July_2010 14] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/15_July_2010&action=edit 15] | [http://2010.igem.org/Korea_U_Seoul/16_July_2010 16] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/17_July_2010&action=edit 17] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/18_July_2010&action=edit 18] |
[http://2010.igem.org/Korea_U_Seoul/19_July_2010 19] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/20_July_2010&action=edit 20] | [http://2010.igem.org/Korea_U_Seoul/21_July_2010 21] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/22_July_2010&action=edit 22] | [http://2010.igem.org/Korea_U_Seoul/23_July_2010 23] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/24_July_2010&action=edit 24] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/25_July_2010&action=edit 25] |
[http://2010.igem.org/Korea_U_Seoul/26_July_2010 26] | [http://2010.igem.org/Korea_U_Seoul/27_July_2010 27] | [http://2010.igem.org/Korea_U_Seoul/28_July_2010 28] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/29_July_2010&action=edit 29] | [http://2010.igem.org/Korea_U_Seoul/30_July_2010 30] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/31_July_2010&action=edit 31] |
|align="center" width="150pt"|
August | ||||||
M | T | W | T | F | S | S |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/1_August_2010&action=edit 1] | ||||||
[http://2010.igem.org/Korea_U_Seoul/2_August_2010 2] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/3_August_2010&action=edit 3] | [http://2010.igem.org/Korea_U_Seoul/4_August_2010 4] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/5_August_2010&action=edit 5] | [http://2010.igem.org/Korea_U_Seoul/6_August_2010 6] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/7_August_2010&action=edit 7] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/8_August_2010&action=edit 8] |
[http://2010.igem.org/Korea_U_Seoul/9_August_2010 9] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/10_August_2010&action=edit 10] | [http://2010.igem.org/Korea_U_Seoul/11_August_2010 11] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/12_August_2010&action=edit 12] | [http://2010.igem.org/Korea_U_Seoul/13_August_2010 13] | [http://2010.igem.org/Korea_U_Seoul/14_August_2010 14] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/15_August_2010&action=edit 15] |
[http://2010.igem.org/Korea_U_Seoul/16_August_2010 16] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/17_August_2010&action=edit 17] | [http://2010.igem.org/Korea_U_Seoul/18_August_2010 18] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/19_August_2010&action=edit 19] | [http://2010.igem.org/Korea_U_Seoul/20_August_2010 20] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/21_August_2010&action=edit 21] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/22_August_2010&action=edit 22] |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/23_August_2010&action=edit 23] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/24_August_2010&action=edit 24] | [http://2010.igem.org/Korea_U_Seoul/25_August_2010 25] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/26_August_2010&action=edit 26] | [http://2010.igem.org/Korea_U_Seoul/27_August_2010 27] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/28_August_2010&action=edit 28] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/29_August_2010&action=edit 29] |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/30_August_2010&action=edit 30] | [http://2010.igem.org/Korea_U_Seoul/31_August_2010 31] |
|align="center" width="150pt"|
September | ||||||
M | T | W | T | F | S | S |
[http://2010.igem.org/Korea_U_Seoul/1_September_2010 1] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/2_September_2010&action=edit 2] | [http://2010.igem.org/Korea_U_Seoul/3_September_2010 3] | [http://2010.igem.org/Korea_U_Seoul/4_September_2010 4] | [http://2010.igem.org/Korea_U_Seoul/5_September_2010 5] | ||
[http://2010.igem.org/Korea_U_Seoul/6_September_2010 6] | [http://2010.igem.org/Korea_U_Seoul/7_September_2010 7] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/8_September_2010&action=edit 8] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/9_September_2010&action=edit 9] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/10_September_2010&action=edit 10] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/11_September_2010&action=edit 11] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/12_September_2010&action=edit 12] |
[http://2010.igem.org/Korea_U_Seoul/13_September_2010 13] | [http://2010.igem.org/Korea_U_Seoul/14_September_2010 14] | [http://2010.igem.org/Korea_U_Seoul/15_September_2010 15] | [http://2010.igem.org/Korea_U_Seoul/16_September_2010 16] | [http://2010.igem.org/Korea_U_Seoul/17_September_2010 17] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/18_September_2010&action=edit 18] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/19_September_2010&action=edit 19] |
[http://2010.igem.org/Korea_U_Seoul/20_September_2010 20] | [http://2010.igem.org/Korea_U_Seoul/21_September_2010 21] | [http://2010.igem.org/Korea_U_Seoul/22_September_2010 22] | [http://2010.igem.org/Korea_U_Seoul/23_September_2010 23] | [http://2010.igem.org/Korea_U_Seoul/24_September_2010 24] | [http://2010.igem.org/Korea_U_Seoul/25_September_2010 25] | [http://2010.igem.org/Korea_U_Seoul/26_September_2010 26] |
[http://2010.igem.org/Korea_U_Seoul/27_September_2010 27] | [http://2010.igem.org/Korea_U_Seoul/28_September_2010 28] | [http://2010.igem.org/Korea_U_Seoul/29_September_2010 29] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/30_September_2010&action=edit 30] |
|align="center" width="150pt"|
October | ||||||
M | T | W | T | F | S | S |
[http://2010.igem.org/Korea_U_Seoul/1_October_2010 1] | [http://2010.igem.org/Korea_U_Seoul/2_October_2010 2] | [http://2010.igem.org/Korea_U_Seoul/3_October_2010 3] | ||||
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/4_October_2010&action=edit 4] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/5_October_2010&action=edit 5] | [http://2010.igem.org/Korea_U_Seoul/6_October_2010 6] | [http://2010.igem.org/Korea_U_Seoul/7_October_2010 7] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/8_October_2010&action=edit 8] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/9_October_2010&action=edit 9] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/10_October_2010&action=edit 10] |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/11_October_2010&action=edit 11] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/12_October_2010&action=edit 12] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/13_October_2010&action=edit 13] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/14_October_2010&action=edit 14] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/15_October_2010&action=edit 15] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/16_October_2010&action=edit 16] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/17_October_2010&action=edit 17] |
[http://2010.igem.org/Korea_U_Seoul/18_October_2010 18] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/19_October_2010&action=edit 19] | [http://2010.igem.org/Korea_U_Seoul/20_October_2010 20] | [http://2010.igem.org/Korea_U_Seoul/21_October_2010 21] | [http://2010.igem.org/Korea_U_Seoul/22_October_2010 22] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/23_October_2010&action=edit 23] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/24_October_2010&action=edit 24] |
[http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/25_October_2010&action=edit 25] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/26_October_2010&action=edit 26] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/27_October_2010&action=edit 27] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/28_October_2010&action=edit 28] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/29_October_2010&action=edit 29] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/30_October_2010&action=edit 30] | [http://2010.igem.org/wiki/index.php?title=Korea_U_Seoul/31_October_2010&action=edit 31] |
Experimental notes
[Discussion] 2010-08-02 ~ 2010-08-29
1. Strategy and overview of iGEM 2010 experiment
2. Design of primers
Primer | Sequence ( 5’ → 3’ ) |
---|---|
PyodA(EcoRI)_F | zz |
mAAA(SpeI)_R | zzzz |
PzntA(XbaI)_F | xzzzz |
RFP(PstI)_R | cxvcxv |
ParsR(XbaI)_F | zzzz |
GFP(PstI)_R | asasas |
[ Preparation of competent cells ] 2010-09-01 ~ 2010-09-03
1. Inoculation of E. coli DH5α and E. coli BL21(DE3) to 3mL LB broth
2. Preparation of 200mL 2x LB broth, TSS solution and LB plates with ampicillin(100μg/mL) and chloramphenicol(25μg /mL), respectively
3. Inoculation of subcultured E. coli to 200mL 2x LB borth
4. Preparation of competent cells by CSBL laboratory protocol
5. Transformation of pUC19 plasmid(10ng/μL) to competent cells for transformation efficiency check
[ Transformation efficiency ] 2010-09-04
Strain | Number of colonies (colonies/μg DNA) |
---|---|
E. coli DH5α | Number of colonies (colonies/μg DNA) |
E. coli BL21(DE3) | 1.5 x 105 |
[ Amplification of BioBrick parts : pSB1A2 and pSB1C3 ] 2010-09-05
1. Confirmed location : pSB1A2-BBa_E0040 (2010 Kit plate 1/ 14K) and pSB1C3-BBa_J04450 (2010 Kit plate 1/ 3A)
2. 20uL suspension by autoclaved distilled water
3. 3uL transformation to E. coli DH5α
4. Plating to LB(Amp100), LB(Cm25)
[ Genomic DNA extraction ] 2010-09-06
1. Inoculation for plasmid DNA purification
2. E. coli K12 genomic DNA extraction by AccuPrep® Genomic DNA Extraction Kit
3. Confirmation of genomic DNA by agarose gel electrophoresis (Figure 1)
4. Quantification of DNA concentration by NanoDrop : 137.5ng/μL
[ Plasmid DNA extraction : pSB1A3 and pSB1C3 ] 2010-09-07
1. Plasmid miniprep by LaboPass™ Plasmid Mini (Plasmid DNA purification kit)
2. Confirmation of extracted plasmids by agarose gel electrophoresis (Figure 2)
3. Quantification of DNA concentration by NanoDrop
[ PCR : promoters and reporter genes ] 2010-09-13 ~ 2010-09-16
1. PCR : PyodA-mAAA, PzntA-RFP(BBa_E1010) and ParsR-GFP(BBa_E0040)
Reagent | Volume (μL) |
---|---|
2.5mM dNTP | 3 |
10x buffer | 5 |
Plasmid template (20ng/μL) | 2 |
Primers (10pmole/μL) | 4 |
α-Taq DNA polymerase (5U/μL) | 0.5 |
D.W. | 35.5/total=50 |
95˚C(2’)-[95˚C(20”)-55˚C(20”)-72˚C(2’)]30-72˚C(5’)-4˚C |
2. Confirmation of PCR products by agarose gel electrophoresis (Figure 3)
3. Purified PCR products
4. Quantification of DNA concentration by NanoDrop
[ Digestion] 2010-09-17
1. Digestion of PCR products and pSB1A2
- 1) PyodA-mAAA : EcoRI and SpeI
- 2) PzntA-RFP : XbaI and PstI
- 3) pSB1A3 : EcoRI and PstI
Reagent | Volume (μL) |
---|---|
DNA (about 30ng/μL) | 30 |
10x NEB buffer 2 | 5 |
BSA (10mg/mL) | 0.5 |
Appropriate 1st and 2nd restriction enzymes | 2 (each 1) |
D.W. | 12.5 / total = 50 |
Completely digestion at 37˚C for 2 hours (at least)
and stop at 80˚C for 20min |
2. Confirmation of digested products by agarose gel electrophoresis (Figure 4)
3. Quantification of DNA concentration by NanoDrop
[Chuseok, Korean thanksgiving day] 2010-09-20 ~ 2010-09
[ Ligation & Transformation ] 2010-09-27
1. Ligation of each parts : PyodA-mAAA, PzntA-RFP and pSB1A2
Reagent | Volume (μL) |
---|---|
10x T4 DNA ligase reaction buffer | 2 |
T4 DNA ligase | 2 |
Each of the digests | 2 + 2 + 2 = 8 |
D.W. | 8 / total = 20 |
Incubation at room temperature for 30min
and stop at 80˚C for 20min |
2. Transformation to E. coli DH5α
[ Confirmation of 1st cloning ] 2010-09-28
1. Check : the color of colonies (pSB1A2 : green, recombinant plasmid : white)
2. Inoculation of white colonies to 3mL LB(Amp100)
[ Plasmid DNA extraction : pSB1A2-( PyodA-mAAA-PzntA-RFP) ] 2010-09-29
1. Plasmid DNA purification by LaboPass™ Plasmid Mini
2. Confirmation of extracted plasmids by agarose gel electrophoresis (Figure 5)
3. Recombinant plasmid sequencing by COSMO GeneTech
[ Digestion ] 2010-10-01 ~ 2010-10-03
1. Check : recombinant plasmid sequence
2. Selection of correct clones
3. Digestion of PCR products(ParsR-GFP) and pSB1C3
- 1) ParsR-GFP : EcoRI and SpeI
- 2) pSB1C3 : EcoRI and PstI
Reagent | Volume (μL) |
---|---|
DNA (about 30ng/μL) | 30 |
10x NEB buffer 2 | 5 |
BSA (10mg/mL) | 0.5 |
Appropriate 1st and 2nd restriction enzymes | 2 (each 1) |
D.W. | 12.5 / total = 50 |
Completely digestion at 37˚C for 2 hours (at least)
and stop at 80˚C for 20min |
4. Confirmation of digested products by agarose gel electrophoresis (Figure 6)
5. Quantification of DNA concentration by NanoDrop
6. Ligation of each parts : PyodA-mAAA-PzntA-RFP, ParsR-GFP and pSB1C3
Reagent | Volume (μL) |
---|---|
10x T4 DNA ligase reaction buffer | 2 |
T4 DNA ligase | 2 |
Each of the digests | 2 + 2 + 2 = 8 |
D.W. | 8 / total = 20 |
Incubation at room temperature for 30min
and stop at 80˚C for 20min |
7. Transformation to E. coli DH5α
[ Confirmation of 2nd cloning ] 2010-10-06
1. Check : the color of colonies (pSB1C3 : red, recombinant plasmid : white)
2. Inoculation of white colonies to 3mL LB(Amp100)
[ Plasmid DNA extraction : pSB1C3-( PyodA-mAAA-PzntA-RFP-ParsR-GFP) ] 2010-10-07
1. Plasmid DNA purification by LaboPass™ Plasmid Mini
2. Confirmation of extracted plasmids by agarose gel electrophoresis (Figure 7)
3. Recombinant plasmid full-sequencing by COSMO GeneTech
[ Completion : Heavy-metal detector ] 2010-10-18
1. Check : recombinant plasmid sequence
2. Selection of correct clones
3. Transformation to E. coli BL21(DE3) for expression test