Aim of the construct

• Chromoproteins represent ideal reporter genes. Unlike fluorescent protein, from which chromoproteins were originally derived, they require no specialised optical equipment for detection. Our aim is to use a selection of such proteins, donated by the iGEM 2012 Uppsala team, as our most basic and main output in regards to the Marchantia framework.
  We decided to test the following chromoprotein constructs:
  
• 35s - eforRed - nosT
• 35s - eforRed - N7 - nosT
• 35s - amilCP - N7 - nosT
• 35s - tsPurple - nosT
• 35s - tsPurple - N7 - nosT
• 35s - asPink - N7 - nosT
• 35s - aeBlue - N7 - nosT

By expressing the chromoproteins in the pGreen vector with the 35s promoter and the nosT terminaor, we control for the possiblity of the promoter/terminator not functioning in our chassis. The N7 fragment allows for the selected constructs to localise chromoprotein to the nucleus. It is believed by Bernardo Pollak that this may end up increasing the likelihood of detecting the chromoproteins with the naked eye.

End goal plasmid

• insert image here

PCR

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• UIDs of primers/plasmids used:
  • A2, P12, P13
  • B3, P14, P13
  • A4, P12, P13
• PCR Spreadsheet (on google drive)
• Gel 27/07/2014

Gibson

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E-Coli transformation

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Agrobacteria transformation

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Spore preparation

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Spore transformation

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Evaluation

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