Paris/September 17

From 2007.igem.org

(Difference between revisions)
Line 25: Line 25:
* '''Quantity:''' ng/µL  <br>
* '''Quantity:''' ng/µL  <br>
* '''Expected size:'''  1403 kb
* '''Expected size:'''  1403 kb
 +
 +
== Digestions ==
 +
 +
{|
 +
|- style="background: #ccccff;"
 +
! colspan="5" style="background: #ccccff;" | Digestion Products
 +
|- style="background: #ccccff; text-align: center;"
 +
|width=5%| Number
 +
|width=24%| Product Name
 +
|width=24%| Matrix Name
 +
|width=5%| Enzyme 1
 +
|width=5%| Enzyme 2
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|width=5%| Size
 +
|width=34%| Description
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|width=1%|
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|width=1%|
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|width=1%|
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|- style="background: #cccccc;" 
 +
| style="background: #FF99FF;" |D56
 +
|araC/pBad>>RBS-Cre BBinsert
 +
|L26.1 araC/pbad>>RBS-Cre in <bbpart>J61002</bbpart>
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|EcoRI
 +
|Pst1
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|
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|araC/pBad>>RBS-Cre BBinsert (for transfer to the low-copy plasmid vector)
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|
 +
|
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|
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|- style="background: #cccccc;" 
 +
| style="background: #FF99FF;" |D66
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|KanR <bbpart>BBa_J61025</bbpart> FV
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|(MP12.1, MP12.2) <bbpart>BBa_P1003</bbpart> in <bbpart>pSB1A2</bbpart>
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|EcoR
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|XbaI
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|
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|KanR <bbpart>BBa_J61025</bbpart> (for forward introduction of a casette for Wanner K insertion)
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|
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|
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|
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|- style="background: #cccccc;" 
 +
| style="background: #ccffcc;" |D71
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|araC/pBad>>lox71-gfptripart FI
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|L35.1 araC/pBad>>lox71-gfptripart <bbpart>BBa_J61002</bbpart>
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|EcoRI
 +
|SpeI
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|
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|araC/pBad>>lox71-gfptripart for use as a FI
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|
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|
 +
|
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|- style="background: #cccccc;" 
 +
| style="background: #ccffcc;" |D72
 +
|<bbpart>pSB2K3</bbpart> BBvector
 +
|MP16 <bbpart>pSB2K3</bbpart>::attP
 +
|EcoRI
 +
|PstI
 +
|
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|Low copy number plasmid, BB vector
 +
|
 +
|
 +
|
 +
|- style="background: #cccccc;" 
 +
| style="background: #ccffcc;" |D73
 +
|<bbpart>pSB2K3</bbpart> BBvector
 +
|MP16 <bbpart>pSB2K3</bbpart>::attP
 +
|EcoRI
 +
|SpeI
 +
|
 +
|Comparison of digestion efficiency between Pst and Spe in NEB2
 +
|
 +
|
 +
|
 +
|- style="background: #cccccc;" 
 +
| style="background: #ccffcc;" |D75
 +
|AraC-pBAD-RBS-CRE FI
 +
|L26.1 AraC-pBAD-RBS-CRE
 +
|EcoRI
 +
|SpeI
 +
|
 +
|FI, Comparison of digestion efficiency between Pst and Spe in NEB2 (see D56)
 +
|
 +
|
 +
|
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|- style="background: #cccccc;" 
 +
| style="background: #ccffcc;" |D76
 +
|DGAT BB
 +
|P14 DGAT
 +
|EcoRI
 +
|PstI
 +
|
 +
|DGAT
 +
|
 +
|
 +
|
 +
|}

Revision as of 14:39, 17 September 2007

DapA Wanner deletion: pKD13 - DapA homologies PCR

  • Material:

- pKD13: 5 µL (bacteria)
- Tampon GC: 10µL
- dNTP: 1µL (final 200µmol each)
- Taq: 0.5µL
- Water: 32.5µL
- Primer1: 33: 2.5µL (final 0.5µM)
- Primer2: 34: 2.5µL (final 0.5µM)

File:WannerDapA09172007.jpg

  • PCR Program:

35 cycles:
- 98°C: 5min
- 98°C: 45s (X35)
- 69°C: 30s (X35)
- 72°C: 1min (X35)
- 72°C: 10 min
- 4°C: inf

  • Quantity: ng/µL
  • Expected size: 1403 kb

Digestions

Digestion Products
Number Product Name Matrix Name Enzyme 1 Enzyme 2 Size Description
D56 araC/pBad>>RBS-Cre BBinsert L26.1 araC/pbad>>RBS-Cre in J61002 EcoRI Pst1 araC/pBad>>RBS-Cre BBinsert (for transfer to the low-copy plasmid vector)
D66 KanR BBa_J61025 FV (MP12.1, MP12.2) BBa_P1003 in pSB1A2 EcoR XbaI KanR BBa_J61025 (for forward introduction of a casette for Wanner K insertion)
D71 araC/pBad>>lox71-gfptripart FI L35.1 araC/pBad>>lox71-gfptripart BBa_J61002 EcoRI SpeI araC/pBad>>lox71-gfptripart for use as a FI
D72 pSB2K3 BBvector MP16 pSB2K3::attP EcoRI PstI Low copy number plasmid, BB vector
D73 pSB2K3 BBvector MP16 pSB2K3::attP EcoRI SpeI Comparison of digestion efficiency between Pst and Spe in NEB2
D75 AraC-pBAD-RBS-CRE FI L26.1 AraC-pBAD-RBS-CRE EcoRI SpeI FI, Comparison of digestion efficiency between Pst and Spe in NEB2 (see D56)
D76 DGAT BB P14 DGAT EcoRI PstI DGAT
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