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Team:Newcastle/11 August 2010
From 2010.igem.org
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Contents |
Gibson cloning of the rocF BioBrick
Aim
The aim of this experiment is to assemble the fragments we extracted yesterday in order to construct the rocF BioBrick and clone it into pSB1C3 in a single step isothermal reaction.
Materials and Protocol
Please refer to: Gibson cloning for materials required and the protocol for the reaction.
Result
Subtilin Immunity BioBrick
Aims
Our aims for today are to run the gel electrophoresis to check whether we have the correct fragement sizes on the four parts that are we amplified yesterday. If the PCR worked, we will then perform gel extraction and then perform another gel electrophoresis for the extracted gel in order to obtain our BioBrick parts.
Materials and protocol
Please refer to the gel electrophoresis and the gel extraction protocols.
Results
Figure #: Gel electrophoresis of the PCR products of the parts required for the subtilin immunity BioBrick.
- Lane 1: 1 kb DNA ladder
- Lane 2: Plasmid Vector (pSB1C3)
- Lane 3: Promoter and RBS (pVeg-SpoVG)
- Lane 4: spaIFEG Gene Cluster
- Lane 5: Double terminator
- Lane 6: 100 bp DNA ladder
Discussion
Plasmid Vector (lane 2), Promoter & RBS (lane 3) and Double terminator (lane 5) showed up. spaIFEG PCR tube (lane 4) did not show up. We think that this occurrence was due to the Tm error (it should be 63°C as opposed to 46°C). Therefore, another PCR and gel electrophoresis were performed.
Transformation of hyperspank and spoVG
Aim
To transform competent E. coli DH5α with hyperspank and spoVG.
Materials and Protocol
Please refer to transformation of E. coli.
Go back to our main Lab book page
   
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