Team:TU Delft/27 July 2010 content

From 2010.igem.org

Alkane degradation

Yesterday's digestion products were checked on a gel.

1% agarose of plasmid check using digestion reaction. Gel runned at 100V for 1 hour. Of all samples 10 μL was loaded with 2 μL loadingbuffer. 5 μL was loaded of marker

Lane description:

As can be seen from the gel, all digestions went well, let's hope that the ligation / transformation also go well!

Transformation

The ligation mixes were incubated overnight. We transformed 10 μL of these ligations in Top10 competent cells, and these were grown on LB-plates with antibiotic over night.

RBS Characterization

For our RBS characterization project, 5 different RBS sequences from the [http://partsregistry.org/Ribosome_Binding_Sites/Prokaryotic/Constitutive/Anderson Anderson RBS family] where placed in front of GFP and measured over 18 hours using a Gen5 fluorenscence and absorbance plate reader

Strength was calculated by taking the mean of the ratio between the expression of known RBS ([http://partsregistry.org/Part:BBa_B0032 B0032]) and expression of Anderson RBS over some time. Expression being the measured fluorescence divided by measured biomass (absorbance, OD).

The measurements where taken from the part of the curve where optimal growth can be assumed, so from 0:40 until 2:30

Next step: We want to relate the RBS sequence to the strength, so it might be possible to say predict something about the other Anderson RBS sequences.