Team:Newcastle/11 August 2010

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=Gibson cloning of the ''rocF'' BioBrick=
 
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==Aim==
 
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The aim of this experiment is to assemble the fragments we extracted [[Team:Newcastle/10_August_2010|yesterday]] in order to construct the [[Team:Newcastle/Urease|''rocF'' BioBrick]] and clone it into pSB1C3 in a single step isothermal reaction.
 
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==Materials and Protocol==
 
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Please refer to: [[Team:Newcastle/Gibson Cloning|Gibson cloning]] for materials required and the protocol for the reaction.
 
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==Result==
 
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The final result about the success of the Gibson reaction will be found on 13th August, 2010.
 
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==Discussion==
 
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We followed the whole procedure perfectly and we would be getting results by 13th August, 2010.
 
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=Subtilin Immunity BioBrick=
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=Gel electrophoresis for the subtilin immunity fragments=
==Aims==
==Aims==
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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 [[Team:Newcastle/10 August 2010#Subtilin_Immunity_BioBrick| 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.
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The aim of the experiment is to check whether we have the correct fragment sizes on the four subtilin fragments that are we have amplified [[Team:Newcastle/10 August 2010#Subtilin_Immunity_BioBrick| yesterday]].
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[[Image:Newcastle_Loaded_Gel_for_Subtilin_Immunity_BioBrick.JPG|right|200px|thumb]]
 
==Materials and protocol==
==Materials and protocol==
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Please refer to the [[Team:Newcastle/Gel_electrophoresis| gel electrophoresis]] and the [[Team:Newcastle/Gel_extraction| gel extraction]] protocols.
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Please refer to the [[Team:Newcastle/Gel_electrophoresis| gel electrophoresis]] protocol.
==Results==
==Results==
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[[Image:Newcastle_Subtilin_Immunity_Gel_1.jpg|300px]]
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[[Image:Newcastle Subtilin Immunity Gel 1b.jpg|300px]]
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'''Figure #''': Gel electrophoresis of the PCR products of the parts required for the subtilin immunity BioBrick.
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'''Figure 1''': Gel electrophoresis of the PCR products of the parts required for the subtilin immunity BioBrick.
* '''Lane 1''': 1 kb DNA ladder
* '''Lane 1''': 1 kb DNA ladder
* '''Lane 2''': Plasmid Vector (pSB1C3)
* '''Lane 2''': Plasmid Vector (pSB1C3)
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==Discussion==
==Discussion==
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Plasmid Vector (lane 2), Promoter & RBS (lane 3) and Double terminator (lane 5) showed up.
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The correct band size were observed for for the Plasmid Vector (lane 2), Promoter & RBS (lane 3) and Double terminator (lane 5). However no band was observed  for the ''spaIFEG''(lane 4).  
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''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.
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=Transformation of hyperspank and spoVG=
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==Aim==
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To transform competent ''E. coli'' DH5α with hyperspank and spoVG.
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==Materials and Protocol==
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Please refer to [[TeamNewcastleTransformation_of_E._coli|transformation of ''E. coli'']].
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==Conclusion==
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No band was observed for the ''spaIFEG''. This could be due to the wrong melting temperature.
'''Go back to our main [[Team:Newcastle/notebook| Lab book]] page'''
'''Go back to our main [[Team:Newcastle/notebook| Lab book]] page'''
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Latest revision as of 00:11, 26 October 2010

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Contents

Gel electrophoresis for the subtilin immunity fragments

Aims

The aim of the experiment is to check whether we have the correct fragment sizes on the four subtilin fragments that are we have amplified yesterday.

Materials and protocol

Please refer to the gel electrophoresis protocol.

Results

Newcastle Subtilin Immunity Gel 1b.jpg

Figure 1: 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

The correct band size were observed for for the Plasmid Vector (lane 2), Promoter & RBS (lane 3) and Double terminator (lane 5). However no band was observed for the spaIFEG(lane 4).

Conclusion

No band was observed for the spaIFEG. This could be due to the wrong melting temperature.

Go back to our main Lab book page

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