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Team:TU Delft/30 July 2010 content
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| - | + | =Alkane Sensing, Solvent Tolerance and Salt Tolerance= | |
''by Pieter'' | ''by Pieter'' | ||
| + | |||
| + | I tried to assemble the remaining biobricks from various sub projects. The goal is to assemble the following: | ||
| + | * AlkS + E0240 | ||
| + | * PalkB + J06702 | ||
| + | * J61101 + bbc1 | ||
| + | * J61101 + PhPFDalpha | ||
| + | * J61101 + PhPFDbeta | ||
| + | To achieve this J06702, bbc1, PhPFDalpha and PhPFDbeta were amplified by PCR. The J61101 was cut open with S and P after which the biobricks were inserted. AlkS was inserted in the backbone with E0240. | ||
| + | |||
| + | ==PCR with PFX== | ||
| + | Per sample the following mix was used: | ||
| + | *1,5 ul 10 mM dNTPs | ||
| + | *5 ul Enhancer | ||
| + | *5 ul 10x PFX Buffer | ||
| + | *1 ul 50 mM MgSO4 | ||
| + | *3 ul 5 pmol/ul Forward Primer | ||
| + | *3 ul 5 pmol/ul Reverse Primer | ||
| + | *0.6 ul PFX Polymerase | ||
| + | *1 ul DNA Template | ||
| + | *29.9 ul H2O | ||
| + | |||
| + | PCR Program: | ||
| + | start: | ||
| + | *2 min 95 C | ||
| + | *1 min 50 C | ||
| + | *1 min 68 C | ||
| + | |||
| + | repeated 25 times: | ||
| + | *1 min 95 C | ||
| + | *1 min 50 C | ||
| + | *1 min 68 C | ||
| + | |||
| + | end: | ||
| + | *10 min 68 C | ||
Revision as of 09:59, 2 August 2010
Alkane Sensing, Solvent Tolerance and Salt Tolerance
by Pieter
I tried to assemble the remaining biobricks from various sub projects. The goal is to assemble the following:
* AlkS + E0240 * PalkB + J06702 * J61101 + bbc1 * J61101 + PhPFDalpha * J61101 + PhPFDbeta
To achieve this J06702, bbc1, PhPFDalpha and PhPFDbeta were amplified by PCR. The J61101 was cut open with S and P after which the biobricks were inserted. AlkS was inserted in the backbone with E0240.
PCR with PFX
Per sample the following mix was used:
- 1,5 ul 10 mM dNTPs
- 5 ul Enhancer
- 5 ul 10x PFX Buffer
- 1 ul 50 mM MgSO4
- 3 ul 5 pmol/ul Forward Primer
- 3 ul 5 pmol/ul Reverse Primer
- 0.6 ul PFX Polymerase
- 1 ul DNA Template
- 29.9 ul H2O
PCR Program: start:
- 2 min 95 C
- 1 min 50 C
- 1 min 68 C
repeated 25 times:
- 1 min 95 C
- 1 min 50 C
- 1 min 68 C
end:
- 10 min 68 C
