Team:TMU-Tokyo/Notebook experiment/4th

From 2013.igem.org

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Get Fragment 1-2<Br>
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Electro Poration<Br>
Electro Poration<Br>
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<b>September 25th</b><Br>
<b>September 25th</b><Br>
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Check PCR.<Br>
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<div style="text-align:center;">
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<Table cellpadding="5">
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<Tr>
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<Td>Fragment</Td><Td>Fragment 3 - ApCm<Br>(control)</Td><Td>Fragment 3 - ApCm</Td>
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</Tr>
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<Tr>
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<Td>Primer</Td><Td>141-49<Br>141-1</Td><Td>CmNR-Not<Br>141-1</Td>
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</Tr>
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<Tr>
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<Td>Templete</Td><Td>(Cell suspention)</Td><Td>(Cell suspention)</Td>
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</Tr>
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</Table>
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</div>
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Result.<Br>
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<IMG Src="https://static.igem.org/mediawiki/2013/8/8b/TMU-Tokyo_0925.jpg"><Br>
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1-7: 141-49 and 141-1, 8: control(141-49 and 141-8),<Br>
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9-15: 141-1 and CmNR-Not, 16: control(141-1 and CmNR-Not)<Br>
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<Br>
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<Br>
<b>September 26th</b><Br>
<b>September 26th</b><Br>
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P1 transduction<Br>
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<Table><Tr><Td>Fragment 3 to Fragment 1-2</Td></Tr></Table>
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Get Fragment 1-2-3.<Br>
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<Br>
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<Br>
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Check PCR<Br>
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<div style="text-align:center;">
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<Table cellpadding="5">
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<Tr>
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<Td>Fragment</Td><Td>Fragment 1-2-3</Td><Td>Fragment 1-2<Br>(control)</Td>
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</Tr>
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<Tr>
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<Td>Primer</Td><Td>CmNR-Not<Br>141-1</Td><Td>141-49<Br>141-1</Td>
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</Tr>
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<Tr>
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<Td>Templete</Td><Td>(Cell suspention)</Td><Td>(Cell suspention)</Td>
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</Tr>
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</Table>
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</div>
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Result.<Br>
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<IMG Src="https://static.igem.org/mediawiki/2013/8/8a/TMU-Tokyo_0926.jpg"><Br>
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1-7: Fragment 1-2-3, 8: cotrol, 9-10: Fragment 1,<Br>
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11-13: Fragment 2, 14-15: Fragment 3 - Ap, 16-17: Fragment 3 - Cm<Br>
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18-19: Fragment 3 - AmCm<Br>
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<Br>
<b>September 27th</b><Br>
<b>September 27th</b><Br>

Revision as of 23:51, 27 September 2013



Experiment > 4th week


September 22th
Mini prep following plasmid.
fimE-pBAD(in pSB1A3), pSB1C3-Km, pSB1A3-Km
Result.

1: pSB1C3-Km, 2-4: pSB1A3-Km, 5-6: fimE-pBAD


Check PCR.
Fragment(control)Fragment 3 - ApCm
Primer141-49
SaApL
141-49
SaApL
TempleteEscherichia coli
MG1655 red
(Cell suspention)
(Cell suspention)
Result.

1-9: Fragment 3 - ApCm

Check PCR.
Fragment(control)Fragment 3 - ApCmFragment 3 - ApCm
Primer2 sets of the others141-1
141-20
141-1
141-74
TempleteEscherichia coli
MG1655 red
(Cell suspention)
(Cell suspention)
Result.

1-7: 141-1 and 141-20, 8: control(141-1 and 141-20),
9-15: 141-1 and 141-74, 16: control(141-1 and 141-74),


Digestion following plasmid.
pSB1C3-Km-Blunt


P1 Preparation following fragment.
Fragment 1, 2


September 23th
PCR following fragment.
FragmentFragment 3 - ApCm
PrimermhpA-CmC6
SaApL
TempleteFragment 3 - Cm
Ap(SaApR-SaApL)
Result.

1: Fragment 3 - Cm, 2: Fragment 3 - ApCm,
3: pSB1C3-Km-Blunt


Check PCR.
FragmentFragment 3 - Ap
(control)
Fragment 3 - ApCm
PrimerCmNR-Not
SaApL
CmNR-Not
SaApL
Templete(Cell suspention)(Cell suspention)
Result.

1-31: Fragment 3 - ApCm


Electro Pration
Fragment 3 - Cm to Escherichia coli MG1655 red.


P1 Preparation
Fragment 3 - ApCm


September 24th
P1 transdunction
Fragment 1, 2
Result.



FragmentFragment 3 - Ap
(control)
Fragment 3 - ApCm
PrimerCmNR-Not
141-1
CmNR-Not
141-1
Templete(Cell suspention)(Cell suspention)
Result.

1-16: Fragment 3 - ApCm


P1 transduction
Fragment 2 to Fragment 1
Fragment 1 to Fragment 2
Result.


Get Fragment 1-2

Electro Poration
Fragment 1, 2, 3 - Ap, 3 - Cm, 3 - ApCm (in pSBC3-Km) to Escherichia coli DH1
Result.



September 25th
Check PCR.
FragmentFragment 3 - ApCm
(control)
Fragment 3 - ApCm
Primer141-49
141-1
CmNR-Not
141-1
Templete(Cell suspention)(Cell suspention)
Result.

1-7: 141-49 and 141-1, 8: control(141-49 and 141-8),
9-15: 141-1 and CmNR-Not, 16: control(141-1 and CmNR-Not)


September 26th
P1 transduction
Fragment 3 to Fragment 1-2
Get Fragment 1-2-3.


Check PCR
FragmentFragment 1-2-3Fragment 1-2
(control)
PrimerCmNR-Not
141-1
141-49
141-1
Templete(Cell suspention)(Cell suspention)
Result.

1-7: Fragment 1-2-3, 8: cotrol, 9-10: Fragment 1,
11-13: Fragment 2, 14-15: Fragment 3 - Ap, 16-17: Fragment 3 - Cm
18-19: Fragment 3 - AmCm

September 27th
September 28th


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