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We designed a (super)plasmid with the code for three different chaperones, GroES/GroEL, DnaK and Trigger factor each with a different promotor. The purpose of this was to be able to induce transcription of the specific chaperones in different combinations. Four other plasmids were designed for the substrates; E-GFP bound to Amyloid-beta or Tau and M-Neongreen protein bound to Amyloid-beta or Tau, for these substrates we used a fourth promotor. After some research, we found that both E-GFP and M-Neongreen are suitable fusion proteins for proteins that are hard to express in E.coli and decided to use both. | We designed a (super)plasmid with the code for three different chaperones, GroES/GroEL, DnaK and Trigger factor each with a different promotor. The purpose of this was to be able to induce transcription of the specific chaperones in different combinations. Four other plasmids were designed for the substrates; E-GFP bound to Amyloid-beta or Tau and M-Neongreen protein bound to Amyloid-beta or Tau, for these substrates we used a fourth promotor. After some research, we found that both E-GFP and M-Neongreen are suitable fusion proteins for proteins that are hard to express in E.coli and decided to use both. | ||
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<br>The plasmid with chaperones and the plasmid with substrates have different antibiotic resistance genes to make it possible to select the bacteria’s that expresses both plasmids. </p> | <br>The plasmid with chaperones and the plasmid with substrates have different antibiotic resistance genes to make it possible to select the bacteria’s that expresses both plasmids. </p> | ||
<p>We will induce the expression of the chaperones before we induce the expression of the substrates to make sure that there are chaperones present when the translation of the substrates begins. </p> | <p>We will induce the expression of the chaperones before we induce the expression of the substrates to make sure that there are chaperones present when the translation of the substrates begins. </p> |
Revision as of 11:42, 27 June 2017