Team:Bielefeld-Germany/Protocols/Overview

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Revision as of 13:03, 20 September 2011

Molecular

In this section of our protocol pages you can read more about our methods for cloning and BioBrick assembly.

Genetic engineering is a basic tool of synthetic biology. With the help of standardized DNA building blocks (BioBricks) it is comparatively easy to create new and modify existing natural systems. The methods we have used in our project to create BioBricks and to modify, mutate, transform and analyse DNA are presented here. Methods used: Electroporation; chemical transformation; Standard, Freiburg, Gibson and 3A BioBrick assembly; restriction analysis; colony PCR; site directed mutagenesis

Production

These are the protocols for the cultivations and the downstream processing.

Before one is able to work with a cell-free system based on biological material, the needed proteins have to be produced and purified first. These methods and the one's we used to characterize BioBricks in vivo are presented here. Used methods: Cultivations in shaking flasks and bioreactor; protein clean-up from medium, periplasm, whole cell and inclusion bodies; UF / DF; IEX; Ni-NTA columns and chromatography; recrystallization and immobilization of S-layer proteins

Analytics

Protocols for the analytical methods we used.

DNA and proteins are very small and cannot be seen by the naked eye. To control the success and the results of your upstream and downstream processes, analytical methods are necessary that give reliable results to make DNA or proteins in any way visible for you. The analytical methods we used in our project can be found here. Used methods: Fluorescence measurement; SDS-PAGE; MALDI-TOF; HPLC; LC-ESI-qTOF-MS/MS; molecular beacons; extraction

Material

Chemicals, enzymes and kits we used in our lab work.

Chemical and biological reactions need defined conditions to work as expected. Read more about chemicals, enzymes, kits, buffers and media we used in our project here.



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