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Team:Newcastle/4 August 2010
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| + | |1st fragment of ''rocF'' CDS | ||
| + | |''B. subtilis'' 168 chromosome | ||
| + | |P1S1 forward | ||
| + | |P2S1 reverse | ||
| + | |58 | ||
| + | |246 approx. | ||
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Revision as of 09:21, 5 August 2010
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Contents |
Cloning the rocF BioBrick
Aim
The aim of today's experiment is to amplify 6 different fragments for the construction of rocF BioBrick with the help of 6 different Phusion PCR.
Materials and Protocol
Please refer to PCR for Phusion PCR protocol. The details for the 6 PCR reactions are mentioned below:
PCR
| Tube | Part to be amplified | Plasmid consisting the part | Forward primer | Reverse Primer | Melting Temperature (Tm in °C) | Size of the fragment (in bp) | Extension time* (in seconds) |
|---|---|---|---|---|---|---|---|
| 1 | Plasmid Vector | pSB1C3 | P1V1 forward | P2V1 reverse | 58 | 2072 approx. | 60 |
| 2 | Pspacoid Promoter | pMutin4 | P1P1 forward | P2P1 reverse | 49 | 106 approx. | 15 |
| 3 | 1st fragment of rocF CDS | B. subtilis 168 chromosome | P1S1 forward | P2S1 reverse | 58 | 246 approx. | 15 |
Gibson assembly
   
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