Team:UT-Tokyo/Sudoku notebook 6

From 2010.igem.org

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{{UT-Tokyo_HEADER}}
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__NOTOC__{{UT-Tokyo_CSS3}}{{UT-Tokyo_CSS_p}}{{UT-Tokyo_Head}}
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<div id="main_all">
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= '''Sudoku''' =
= '''Sudoku''' =
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<html>
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[https://2010.igem.org/Team:UT-Tokyo/Sudoku_abstract  Abstract]      
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<div>
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[https://2010.igem.org/Team:UT-Tokyo/Sudoku_construct Construct]     
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        <ul id="inpagemenu">
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[https://2010.igem.org/Team:UT-Tokyo/Sudoku_experiment Experiment]     
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                <li><a href="/Team:UT-Tokyo/Sudoku_abstract" id="abstract">Introduction</a></li>
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Notebook      
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                <li><a href="/Team:UT-Tokyo/Sudoku_construct" id="construct">System</a></li>
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[https://2010.igem.org/Team:UT-Tokyo/Sudoku_result Result]
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                <li><a href="/Team:UT-Tokyo/Sudoku_modeling" id="modeling">Modeling</a></li>
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                <li><span>Experiments</span></li>
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                <li><a href="/Team:UT-Tokyo/Sudoku_perspective" id="perspective">Perspective</a></li>
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            <li><a href="/Team:UT-Tokyo/Sudoku_reference" id="reference">Reference</a></li>
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            </ul>
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</div>
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<div id="clear"></div>
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</html><br />
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Notebook =>
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[
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June     /
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[https://2010.igem.org/Team:UT-Tokyo/Sudoku_notebook July]    /
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[https://2010.igem.org/Team:UT-Tokyo/Sudoku_notebook_2 August]    /      
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[https://2010.igem.org/Team:UT-Tokyo/Sudoku_notebook_8 September - October]
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]
== '''Notebook''' ==
== '''Notebook''' ==
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====6/25====
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====6/23====
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'''Enxyme digestion'''
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'''Transformation'''
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plasmid 1~7, 9 & 10
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2010 Kit plate1: 6N,1H,5D,17J,4H,18A
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1: 117.2 ng/uL -> 2.56 uL / SP (3 uL)
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2010 Kit plate2: 24C,2K
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2:  59.3 ng/uL -> 5.06 uL / SP (3 uL)
 
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3: 124.2 ng/uL -> 2.42 uL / XP (3 uL)
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====6/24====
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4: 114.5 ng/uL -> 2.62 uL / EX (3 uL)
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'''Inoculation'''
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5: 81.3 ng/uL -> 3.69 uL / SP (3 uL)
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2010 Kit plate1: 6N,1H,5D,17J,4H,18A
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6: 101.7 ng/uL -> 2.95 uL / ES (3 uL)
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2010 Kit plate2: 24C,2K
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7:  81.5 ng/uL -> 3.68 uL / EX (3 uL)
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(6/23 Transformation)
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9:  85.4 ng/uL -> 3.51 uL / SP (3 uL)
 
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10: 73.5 ng/uL -> 4.08 uL / SP (3 uL)
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====6/25====
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====6/28====
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'''Miniprep'''
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====6/29====
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2010 Kit plate1: 6N,1H,5D,17J,4H,18A
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====6/30====
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2010 Kit plate2: 24C,2K
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== Pre-experiment ==
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(6/24 Inoculation)
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=== Make construct ===
 
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==== 7/1 ====
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====6/27====
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'''Transformation'''
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'''Enxyme digestion'''
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2-3 plasmid (6/30 ligation)
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1~7, 9 & 10 plasmid
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< 6/30 transformation → fail >
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1: 117.2 ng/uL -> 2.56 uL / SP
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2:  59.3 ng/uL -> 5.06 uL / SP
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'''Inoculation'''
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3: 124.2 ng/uL -> 2.42 uL / XP
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11 plasmid (6/30 thaw, transformation)
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4: 114.5 ng/uL -> 2.62 uL / EX
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5:  81.3 ng/uL -> 3.69 uL / SP
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'''Enzyme digestion'''
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6: 101.7 ng/uL -> 2.95 uL / ES
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2, 3, 5, 7 plasmid (6/25 miniprep)
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7:  81.5 ng/uL -> 3.68 uL / EX
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2 : 53.9 ng/uL → about 15 uL (shortage) / H / SP (2 uL)
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9: 85.4 ng/uL -> 3.51 uL / SP
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3 : 124.2 ng/uL → 8 uL / M / XP (2 uL)
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10: 73.5 ng/uL -> 4.08 uL / SP
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5 : 81.3 ng/uL → 12.3 uL / H / SP (2 uL)
 
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7 : 85.4 ng/uL → 11.7 uL / H / SP (2 uL)
 
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====6/28====
'''Gel extract'''
'''Gel extract'''
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2, 3, 5, 7 plasmid (7/1 enzyme cut)
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1~7, 9 & 10 plasmid (6/27 enzyme digestion)
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Label : 100701 数X (2,3,5,9) ゲル抽出
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6 -> failed, others -> succeeded
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====6/29====
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==== 7/2 ====
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'''Cutcheck'''
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'''Miniprep'''
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EScut: 1~5, 7, 9 & 10 (6/27 enzyme digestion)
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2-3, 11 plasmid (7/1 inoculation)
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HindIII cut: 6
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2-3 → 105 ng/uL
 
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11 (1st) → 191.3 ng/uL
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====6/30====
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11 (2nd) → 49.5 ng/uL
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'''Enzyme digestion'''
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2-3, 11 plasmid (7/2 miniprep)
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2-3 : 9.5 uL / H / ES (1.5 uL)
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11 (1st) : 5.2 uL / H /  ES (1.5 uL) → Judged failure by cut check → miniprep again
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11 (2nd) : 20.2 uL / H /  ES (1.5 uL)
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==== 7/3 ====
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'''Gel extract'''
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2-3, 11 plasmid (7/2 enzyme cut)
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Label : 100703 X ゲル抽出
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==== 7/5 ====
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'''Ligation'''
'''Ligation'''
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2-3 (insert, 7/3 ES enzyme cut) & 4 (vector, probably 6/28 gel extract, EX enzyme cut) / 11 (insert; 7/3 ES enzyme cut) & 7 (vector, probably 6/28 gel extract, EX enzyme cut)
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2-3  
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2-3 : 37.5 ng/uL → 5 uL & 4 : 5.2 ng/uL → 5 uL
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2: 5uL, 3: 5uL, ligation mix: 10uL
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11 : 4.2 ng/uL → 7.5 uL & 7 : Unknown → 2.5 uL
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(6/27 Enzyme digestion)
'''Transformation'''
'''Transformation'''
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2-3-4 plasmid (7/5 ligation) / 11-7 plasmid (7/5 ligation)
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2010 Kit plate1: 1N (TetA(C) (Backwards))
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2010 Kit plate2: 22P (OmpA)
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2-3 (6/30 Ligation)
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==== 7/6 ====
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{{UT-Tokyo_Foot}}
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'''Inoculation'''
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5 plasmid (6/23 plate)
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'''Enzyme digestion'''
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1, 4 (), 11 plasmid (7/6 miniprep)
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2-3 : 9.5 uL / H / ES (1.5 uL)
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11 (1st) : 5.2 uL / H /  ES (1.5 uL) → Judged failure by cut check → miniprep again
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11 (2nd) : 20.2 uL / H /  ES (1.5 uL)
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=== Check test ===
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== Main construct ==
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=== Research of SP6 RNApol cDNA and F plasmid ===
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=== Convert lox71(rev) to lox66(rev) ===
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=== Make Hin(rev) ===
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=== Revise MS2 gene ===
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=== Make location(rev)-rbs(rev)-location(rev) ===
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</div>
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{{UT-Tokyo_FOOTER}}
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Latest revision as of 03:00, 28 October 2010

UT-Tokyo

Sudoku


Notebook => [ June / July / August / September - October ]

Notebook

6/23

Transformation

2010 Kit plate1: 6N,1H,5D,17J,4H,18A

2010 Kit plate2: 24C,2K


6/24

Inoculation

2010 Kit plate1: 6N,1H,5D,17J,4H,18A

2010 Kit plate2: 24C,2K

(6/23 Transformation)


6/25

Miniprep

2010 Kit plate1: 6N,1H,5D,17J,4H,18A

2010 Kit plate2: 24C,2K

(6/24 Inoculation)


6/27

Enxyme digestion

1~7, 9 & 10 plasmid

1: 117.2 ng/uL -> 2.56 uL / SP

2: 59.3 ng/uL -> 5.06 uL / SP

3: 124.2 ng/uL -> 2.42 uL / XP

4: 114.5 ng/uL -> 2.62 uL / EX

5: 81.3 ng/uL -> 3.69 uL / SP

6: 101.7 ng/uL -> 2.95 uL / ES

7: 81.5 ng/uL -> 3.68 uL / EX

9: 85.4 ng/uL -> 3.51 uL / SP

10: 73.5 ng/uL -> 4.08 uL / SP


6/28

Gel extract

1~7, 9 & 10 plasmid (6/27 enzyme digestion)

6 -> failed, others -> succeeded


6/29

Cutcheck

EScut: 1~5, 7, 9 & 10 (6/27 enzyme digestion)

HindIII cut: 6


6/30

Ligation

2-3

2: 5uL, 3: 5uL, ligation mix: 10uL

(6/27 Enzyme digestion)


Transformation

2010 Kit plate1: 1N (TetA(C) (Backwards))

2010 Kit plate2: 22P (OmpA)

2-3 (6/30 Ligation)

Retrieved from "http://2010.igem.org/Team:UT-Tokyo/Sudoku_notebook_6"