"
Team:Cambridge/Bioluminescence/Colour
From 2010.igem.org
(Difference between revisions)
| Line 1: | Line 1: | ||
{{:Team:Cambridge/Templates/headerMinimalprototype}}{{:Team:Cambridge/Templates/headerbar|colour=#96d446|linkcolour=#6bbe00|title=Project Firefly: Coloured outputs}} | {{:Team:Cambridge/Templates/headerMinimalprototype}}{{:Team:Cambridge/Templates/headerbar|colour=#96d446|linkcolour=#6bbe00|title=Project Firefly: Coloured outputs}} | ||
smith and smith reported point mutations that change the colour of luciola cruciata luciferase. Different coloured outputs would allow co-reporter assays and | smith and smith reported point mutations that change the colour of luciola cruciata luciferase. Different coloured outputs would allow co-reporter assays and | ||
| - | {{:Team:Cambridge/Templates/RightImage|image=Cambridge- | + | {{:Team:Cambridge/Templates/RightImage|image=Cambridge-Mutagenesis.jpg|caption=Comparison of the colour of Luciola cruciata we ordered from DNA 2.0 and it's colour after a single amino acid substitution}} |
==Methods== | ==Methods== | ||
Revision as of 18:03, 26 October 2010
Project Firefly: Coloured outputs
smith and smith reported point mutations that change the colour of luciola cruciata luciferase. Different coloured outputs would allow co-reporter assays and
Comparison of the colour of Luciola cruciata we ordered from DNA 2.0 and it's colour after a single amino acid substitution
Methods
We used site directed mutagenesis to produce these specific mutations. pSB1C3 with the luciferase and LRE genes is a sizeable plasmid so we performed two separate PCR reactions to generate two halves of the plasmid (using oligonucleotides with the point mutation). PCR products were run on an agarose gel and the relevant bands gel extracted then joined by Gibson Assembly. Cells were transformed with the product. This method avoids cells being transformed with the non-mutated, template plasmid. As an additional check colonies which grew from sucessfully transformed cells were imaged inside a dark box after addition of luciferin to check the colour of the light produced was as expected.
A co-reporter system
Results
We produced...
