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Team:CBNU-Korea/Notebook
From 2010.igem.org
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| - | + | <p>5/7<br> | |
| - | <p>5/7<br | + | <strong>Membership meeting</strong><br> |
| - | <strong>Membership meeting</strong><br | + | New participant of CBNU-KOREA team.<br> |
| - | New participant of CBNU-KOREA team.<br | + | Introduced what is the iGEM, what we should do if we join the competition.<br> |
| - | Introduced what is the iGEM, what we should do if we join the competition.<br | + | 5/15<br> |
| - | 5/15<br | + | <strong>Concept meeting</strong><br> |
| - | <strong>Concept meeting</strong><br | + | Decided 2010 CBNU-KOREA iGEM team’s project(almost 4hr)<br> |
| - | Decided 2010 CBNU-KOREA iGEM team’s project(almost 4hr)<br | + | 5/18<br> |
| - | 5/18<br | + | <strong>Researching </strong><br> |
| - | <strong>Researching </strong><br | + | Finding related articles, paper and so on.<br> |
| - | Finding related articles, paper and so on.<br | + | Essential gene data search<br> |
| - | Essential gene data search<br | + | 5/25<br> |
| - | 5/25<br | + | <strong>Strategy about database</strong><br> |
| - | <strong>Strategy about database</strong><br | + | Planned how to re-construct essential gene database.<br> |
| - | Planned how to re-construct essential gene database.<br | + | 5/27<br> |
| - | 5/27<br | + | <strong>Gathering</strong><br> |
| - | <strong>Gathering</strong><br | + | Gathering the essential genes data from DEG, NCBI.<br> |
| - | Gathering the essential genes data from DEG, NCBI.<br | + | Starting to lean about experimental technique.<br> |
| - | Starting to lean about experimental technique.</ | + | 6/1<br> |
| - | < | + | Clean up the lab room. Too dirty…<br> |
| - | <strong>Plasmid Culture</strong> <br | + | Computer engineering department meeting about project<br> |
| - | · Pick up a single colony from fresh cultured LB agar plate.<br | + | 6/24<br> |
| - | · Our established media and inoculate the cell into the 5ml of fresh LB liquid media (contains antibiotics) and our established media at 37℃ with shaking for 12hr.<br | + | Summer iGEM Workshop Asia <br> |
| - | · plasmid : pSB1C3, pSB1A3, pSB1T3, pSB1K3 (contain J54450)<br | + | 7/19<br> |
| - | 7/20<br | + | <strong>Plasmid Culture</strong> <br> |
| - | <strong>Plasmid extract</strong> <br | + | · Pick up a single colony from fresh cultured LB agar plate.<br> |
| - | · Uses kit and extracted plasmid. (pSB1C3, pSB1A3, pSB1T3, pSB1K3)<br | + | · Our established media and inoculate the cell into the 5ml of fresh LB liquid media (contains antibiotics) and our established media at 37℃ with shaking for 12hr.<br> |
| - | · Confirmed size by electrophoresis.<br | + | · plasmid : pSB1C3, pSB1A3, pSB1T3, pSB1K3 (contain J54450)<br> |
| - | 7/23<br | + | 7/20<br> |
| - | <strong>Plasmid Digest</strong> <br | + | <strong>Plasmid extract</strong> <br> |
| - | · Digest of 4 plasmid.<br | + | · Uses kit and extracted plasmid. (pSB1C3, pSB1A3, pSB1T3, pSB1K3)<br> |
| - | · Used EcoRⅠ-PstⅠ restriction endonuclease.<br | + | · Confirmed size by electrophoresis.<br> |
| - | · Confirmed size by electrophoresis.<br | + | 7/23<br> |
| - | 7/24<br | + | <strong>Plasmid Digest</strong> <br> |
| - | <strong>Primer Design </strong> <br | + | · Digest of 4 plasmid.<br> |
| - | · Designed <em>Vibrio cholerae</em>'s <em>oriC2vc, parA, parB, parS, dif</em> gene primer using ‘Ginko primer’.<br | + | · Used EcoRⅠ-PstⅠ restriction endonuclease.<br> |
| - | 7/28<br | + | · Confirmed size by electrophoresis.<br> |
| - | <strong>DNA Amplification</strong> <br | + | 7/24<br> |
| - | · Amplified <em>ori, parA, parB, parS, dif</em> gene of <em>vibrio cholerae</em>.<br | + | <strong>Primer Design </strong> <br> |
| - | · Confirmed size by electrophoresis.<br | + | · Designed <em>Vibrio cholerae</em>'s <em>oriC2vc, parA, parB, parS, dif</em> gene primer using ‘Ginko primer’.<br> |
| - | <strong>Concentration</strong> <br | + | 7/28<br> |
| - | · Because concentration of <em>parS, dif</em> gene was low, it was hard to confirm result by electrophoresis.<br | + | <strong>DNA Amplification</strong> <br> |
| - | · Concentrated gene sample<br | + | · Amplified <em>ori, parA, parB, parS, dif</em> gene of <em>vibrio cholerae</em>.<br> |
| - | 7/29<br | + | · Confirmed size by electrophoresis. And that’s correct.<br> |
| - | <strong>DNA(gene) Digest</strong> <br | + | <strong>Concentration</strong> <br> |
| - | · Digest of 4 plasmid.<br | + | · Because concentration of <em>parS, dif</em> gene was low, it was hard to confirm result by electrophoresis.<br> |
| - | · Used EcoRⅠ-PstⅠ restriction endonuclease.<br | + | · Concentrated gene sample<br> |
| - | · Confirmed size by electrophoresis.<br | + | 7/29<br> |
| - | 7/30<br | + | <strong>DNA(gene) Digest</strong> <br> |
| - | <strong>Prepared Compentent cell</strong> <br | + | · Digest of 4 plasmid.<br> |
| - | · <em>E.coli</em> DH10B, DH5α, JM109<br | + | · Used EcoRⅠ-PstⅠ restriction endonuclease.<br> |
| - | 8/2<br | + | · Confirmed size by electrophoresis.<br> |
| - | <strong>DNA Ligation/Transformation</strong> <br | + | 7/30<br> |
| - | · Ligation inserting gene in vector<br | + | <strong>Prepared Compentent cell</strong> <br> |
| - | (<em>parA</em>+pSB1A3, <em>parS</em>+pSB1C3, <em>ori</em>+pSB1K3, <em>dif</em>+pSB1A3)<br | + | · <em>E.coli</em> DH10B, DH5α, JM109<br> |
| - | · Transformation (Competent cell : <em>E.coli </em>DH10B)<br | + | 8/2<br> |
| - | · Spreading on plate (each antibiotics contained LB agar media)<br | + | <strong>DNA Ligation/Transformation</strong> <br> |
| - | 8/3<br | + | · Ligation inserting gene in vector<br> |
| - | <strong>Cell Culture</strong> <br | + | (<em>parA</em>+pSB1A3, <em>parS</em>+pSB1C3, <em>ori</em>+pSB1K3, <em>dif</em>+pSB1A3)<br> |
| - | · Pick up a single colony from fresh cultured LB agar plate.<br | + | · Transformation (Competent cell : <em>E.coli </em>DH10B)<br> |
| - | · Our established media and inoculate the cell into the 5ml of fresh LB liquid media (contains antibiotics) and our established media at 37℃ with shaking for 12hr.<br | + | · Spreading on plate (each antibiotics contained LB agar media)<br> |
| - | · vector : <em>parA</em>+pSB1A3, <em>parS</em>+pSB1C3, <em>ori</em>+pSB1K3, <em>dif</em>+pSB1A3<br | + | 8/3<br> |
| - | 8/4<br | + | <strong>Cell Culture</strong> <br> |
| - | <strong>Vector extract</strong> <br | + | · Pick up a single colony from fresh cultured LB agar plate.<br> |
| - | · Uses kit and extracted plasmid.<br | + | · Our established media and inoculate the cell into the 5ml of fresh LB liquid media (contains antibiotics) and our established media at 37℃ with shaking for 12hr.<br> |
| - | (parA+pSB1A3, parS+pSB1C3, ori+pSB1K3, dif+pSB1A3)<br | + | · vector : <em>parA</em>+pSB1A3, <em>parS</em>+pSB1C3, <em>ori</em>+pSB1K3, <em>dif</em>+pSB1A3<br> |
| - | · Confirmed size by electrophoresis.<br | + | 8/4<br> |
| - | 8/9<br | + | <strong>Vector extract</strong> <br> |
| - | <strong>Plasmid Digest</strong> <br | + | · Uses kit and extracted plasmid.<br> |
| - | · Digest of 4 vector.<br | + | (parA+pSB1A3, parS+pSB1C3, ori+pSB1K3, dif+pSB1A3)<br> |
| - | · ori+pSB1K3, parA+pSB1A3, parS+pSB1C3 digests by EcoRⅠ-SpeⅠ restriction endonuclease and dif+pSB1A3 gene digested by XbaⅠ-PstⅠ restriction endonuclease.<br | + | · Confirmed size by electrophoresis.<br> |
| - | · Confirmed size by electrophoresis.<br | + | 8/9<br> |
| - | · Came out as size of <em>ori</em>+pSB1K3 gene is smaller.<br | + | <strong>Plasmid Digest</strong> <br> |
| - | 8/10<br | + | · Digest of 4 vector.<br> |
| - | <strong>Sequencing Order</strong> <br | + | · ori+pSB1K3, parA+pSB1A3, parS+pSB1C3 digests by EcoRⅠ-SpeⅠ restriction endonuclease and dif+pSB1A3 gene digested by XbaⅠ-PstⅠ restriction endonuclease.<br> |
| - | · Analyzed plasmid DNA extraction (cultured cell)<br | + | · Confirmed size by electrophoresis.<br> |
| - | 8/16<br | + | · Came out as size of <em>ori</em>+pSB1K3 gene is smaller.<br> |
| - | <strong>Primer Again design </strong> <br | + | 8/10<br> |
| - | · According to analysis result, primer of <em>ori</em> gene knew wrong fact.<br | + | <strong>Sequencing Order</strong> <br> |
| - | · Designed primer of <em>ori</em> gene again.<br | + | · Analyzed plasmid DNA extraction (cultured cell)<br> |
| - | · Primer of I51020 and p1003 designed.<br | + | 8/16<br> |
| - | 8/20<br | + | <strong>Primer Again design </strong> <br> |
| - | <strong>DNA Amplification</strong> <br | + | · According to analysis result, primer of <em>ori</em> gene knew wrong fact.<br> |
| - | · Amplified I51020, p1003<br | + | · Designed primer of <em>ori</em> gene again.<br> |
| - | · Confirmed size by electrophoresis.<br | + | · Primer of I51020 and p1003 designed.<br> |
| - | 8/23<br | + | 8/20<br> |
| - | <strong>Plasmid Digest</strong> <br | + | <strong>DNA Amplification</strong> <br> |
| - | · I51020 digests by EcoRⅠ-PstⅠ restriction endonuclease.<br | + | · Amplified I51020, p1003<br> |
| - | · p1003 digests by XbaⅠ-PstⅠ restriction endonuclease.<br | + | · Confirmed size by electrophoresis.<br> |
| - | · Confirmed size by electrophoresis.<br | + | 8/23<br> |
| - | 8/24<br | + | <strong>Plasmid Digest</strong> <br> |
| - | <strong>DNA Ligation/Transformation</strong> <br | + | · I51020 digests by EcoRⅠ-PstⅠ restriction endonuclease.<br> |
| - | · Ligated inserting gene in vector<br | + | · p1003 digests by XbaⅠ-PstⅠ restriction endonuclease.<br> |
| - | (Inserted <em>ori</em> gene and p1003 antibiotic resistance cassette in I51020 vector, inserted <em>parS </em>gene and <em>dif </em>gene in pSB1A3 vector)<br | + | · Confirmed size by electrophoresis.<br> |
| - | · Transformation (Competent cell : <em>E.coli </em>DH10B)<br | + | 8/24<br> |
| + | <strong>DNA Ligation/Transformation</strong> <br> | ||
| + | · Ligated inserting gene in vector<br> | ||
| + | (Inserted <em>ori</em> gene and p1003 antibiotic resistance cassette in I51020 vector, inserted <em>parS </em>gene and <em>dif </em>gene in pSB1A3 vector)<br> | ||
| + | · Transformation (Competent cell : <em>E.coli </em>DH10B)<br> | ||
· Spreading on plate (each antibiotics contained LB agar media)</p> | · Spreading on plate (each antibiotics contained LB agar media)</p> | ||
| + | <p>8/25<br> | ||
| + | I51020 transformation failed<br> | ||
| + | Plan to design Cn vector <br> | ||
| + | Ready to P1003, <em>ori</em> miniprep </p> | ||
| + | <p>9/2</p> | ||
| + | <p>Primer design for Cn vector. May be it won’t work….</p> | ||
| + | <p>All parts re-transformation for selection and miniprep.</p> | ||
| + | <p><strong> </strong></p> | ||
| + | <p>9/7 </p> | ||
| + | <p>Cn vector primer arrived.<br> | ||
| + | Ready to PSB1C3 plasmid miniprep<br> | ||
| + | <em>Ori</em> X/S digestion<strong>. </strong></p> | ||
| + | <p><strong> </strong></p> | ||
| + | <p><strong>9/10</strong></p> | ||
| + | <p><strong>Cn vector PCR, purification, ligation.</strong><br> | ||
| + | <strong>Cn vector PCR Product electroporhesis.</strong><br> | ||
| + | <strong>9/15</strong><br> | ||
| + | <strong>Cn vector transformation failed.</strong><br> | ||
| + | <strong>Cn vercor PCR condition re arrangement</strong><br> | ||
| + | <strong>PrctB promoter primer design</strong></p> | ||
| + | <p><strong> </strong></p> | ||
| + | <p><strong>9/25</strong><br> | ||
| + | <strong>Cn vector + <em>or</em>i(X-S) ligation, transformation.</strong><br> | ||
| + | <strong>PrctB primer arrived , PCR </strong></p> | ||
| + | <p><strong>9/27</strong><br> | ||
| + | <strong>Cn vector colony appear. But not red, may be failure.</strong><br> | ||
| + | <strong>PrctB + rctB ligation.</strong><br> | ||
| + | <strong>Cn vector transformation</strong></p> | ||
| + | <p><strong>9/30 </strong><br> | ||
| + | <strong>Cn vector colony appear again. Not red too. Ready to miniprep.</strong><br> | ||
| + | <strong>parB + GFP fusion </strong></p> | ||
| + | <p><strong>10/3</strong><br> | ||
| + | <strong>parA, parS, dif ligation PSB1C3 and PSB1A3</strong><br> | ||
| + | <strong>computer engineering student meeting,</strong></p> | ||
| + | <p><strong> </strong></p> | ||
| + | <p><strong>10/8</strong><br> | ||
| + | <strong>Ori PCR and miniprep</strong><br> | ||
| + | <strong>Ori digestion EP/XP/XS</strong></p> | ||
| + | <p><strong> </strong></p> | ||
| + | <p><strong>10/10</strong></p> | ||
| + | <p><strong>parS, dif parts complete.</strong><br> | ||
| + | <strong>parA re PCR.</strong></p> | ||
| + | <p><strong>10/15</strong><br> | ||
| + | <strong>All members of CBNU-KOREA team meeting</strong></p> | ||
| + | <p><strong>10/21</strong><br> | ||
| + | <strong>Parts making start.</strong></p> | ||
| + | <p><strong>Ori function is not checked.</strong></p> | ||
| + | <p><strong>10/25</strong></p> | ||
| + | <p><strong>Parts sending </strong><br> | ||
| + | <strong>Prepare Team wiki freeze</strong></p> | ||
Latest revision as of 03:22, 28 October 2010
5/7
Membership meeting
New participant of CBNU-KOREA team.
Introduced what is the iGEM, what we should do if we join the competition.
5/15
Concept meeting
Decided 2010 CBNU-KOREA iGEM team’s project(almost 4hr)
5/18
Researching
Finding related articles, paper and so on.
Essential gene data search
5/25
Strategy about database
Planned how to re-construct essential gene database.
5/27
Gathering
Gathering the essential genes data from DEG, NCBI.
Starting to lean about experimental technique.
6/1
Clean up the lab room. Too dirty…
Computer engineering department meeting about project
6/24
Summer iGEM Workshop Asia
7/19
Plasmid Culture
· Pick up a single colony from fresh cultured LB agar plate.
· Our established media and inoculate the cell into the 5ml of fresh LB liquid media (contains antibiotics) and our established media at 37℃ with shaking for 12hr.
· plasmid : pSB1C3, pSB1A3, pSB1T3, pSB1K3 (contain J54450)
7/20
Plasmid extract
· Uses kit and extracted plasmid. (pSB1C3, pSB1A3, pSB1T3, pSB1K3)
· Confirmed size by electrophoresis.
7/23
Plasmid Digest
· Digest of 4 plasmid.
· Used EcoRⅠ-PstⅠ restriction endonuclease.
· Confirmed size by electrophoresis.
7/24
Primer Design
· Designed Vibrio cholerae's oriC2vc, parA, parB, parS, dif gene primer using ‘Ginko primer’.
7/28
DNA Amplification
· Amplified ori, parA, parB, parS, dif gene of vibrio cholerae.
· Confirmed size by electrophoresis. And that’s correct.
Concentration
· Because concentration of parS, dif gene was low, it was hard to confirm result by electrophoresis.
· Concentrated gene sample
7/29
DNA(gene) Digest
· Digest of 4 plasmid.
· Used EcoRⅠ-PstⅠ restriction endonuclease.
· Confirmed size by electrophoresis.
7/30
Prepared Compentent cell
· E.coli DH10B, DH5α, JM109
8/2
DNA Ligation/Transformation
· Ligation inserting gene in vector
(parA+pSB1A3, parS+pSB1C3, ori+pSB1K3, dif+pSB1A3)
· Transformation (Competent cell : E.coli DH10B)
· Spreading on plate (each antibiotics contained LB agar media)
8/3
Cell Culture
· Pick up a single colony from fresh cultured LB agar plate.
· Our established media and inoculate the cell into the 5ml of fresh LB liquid media (contains antibiotics) and our established media at 37℃ with shaking for 12hr.
· vector : parA+pSB1A3, parS+pSB1C3, ori+pSB1K3, dif+pSB1A3
8/4
Vector extract
· Uses kit and extracted plasmid.
(parA+pSB1A3, parS+pSB1C3, ori+pSB1K3, dif+pSB1A3)
· Confirmed size by electrophoresis.
8/9
Plasmid Digest
· Digest of 4 vector.
· ori+pSB1K3, parA+pSB1A3, parS+pSB1C3 digests by EcoRⅠ-SpeⅠ restriction endonuclease and dif+pSB1A3 gene digested by XbaⅠ-PstⅠ restriction endonuclease.
· Confirmed size by electrophoresis.
· Came out as size of ori+pSB1K3 gene is smaller.
8/10
Sequencing Order
· Analyzed plasmid DNA extraction (cultured cell)
8/16
Primer Again design
· According to analysis result, primer of ori gene knew wrong fact.
· Designed primer of ori gene again.
· Primer of I51020 and p1003 designed.
8/20
DNA Amplification
· Amplified I51020, p1003
· Confirmed size by electrophoresis.
8/23
Plasmid Digest
· I51020 digests by EcoRⅠ-PstⅠ restriction endonuclease.
· p1003 digests by XbaⅠ-PstⅠ restriction endonuclease.
· Confirmed size by electrophoresis.
8/24
DNA Ligation/Transformation
· Ligated inserting gene in vector
(Inserted ori gene and p1003 antibiotic resistance cassette in I51020 vector, inserted parS gene and dif gene in pSB1A3 vector)
· Transformation (Competent cell : E.coli DH10B)
· Spreading on plate (each antibiotics contained LB agar media)
8/25
I51020 transformation failed
Plan to design Cn vector
Ready to P1003, ori miniprep
9/2
Primer design for Cn vector. May be it won’t work….
All parts re-transformation for selection and miniprep.
9/7
Cn vector primer arrived.
Ready to PSB1C3 plasmid miniprep
Ori X/S digestion.
9/10
Cn vector PCR, purification, ligation.
Cn vector PCR Product electroporhesis.
9/15
Cn vector transformation failed.
Cn vercor PCR condition re arrangement
PrctB promoter primer design
9/25
Cn vector + ori(X-S) ligation, transformation.
PrctB primer arrived , PCR
9/27
Cn vector colony appear. But not red, may be failure.
PrctB + rctB ligation.
Cn vector transformation
9/30
Cn vector colony appear again. Not red too. Ready to miniprep.
parB + GFP fusion
10/3
parA, parS, dif ligation PSB1C3 and PSB1A3
computer engineering student meeting,
10/8
Ori PCR and miniprep
Ori digestion EP/XP/XS
10/10
parS, dif parts complete.
parA re PCR.
10/15
All members of CBNU-KOREA team meeting
10/21
Parts making start.
Ori function is not checked.
10/25
Parts sending
Prepare Team wiki freeze
