< IGEM:IMPERIAL | 2008 | Prototype | Wetlab
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|1.1 Characterisation of Constitutive promoter and RBS |
Aim : To characterise the different combinations of constitutive promoters and RBS in terms of steady-state protein production. At the end of this testing we will have decided upon a constitutive promoter and RBS combination to use in later test constructs. The combinations of Promoter and RBS are listed below,
|2.1 Characterisation of inducible promoters and RBS |
Aim: To characterise the different combinations of inducible promoters and RBS in terms of response time, transfer function and maximum steady-state level of protein. Each of these constructs required the use of the constitutive promoter-RBS to express the transcriptional regulators. By the end of this testing, our extensive characterisation of the different combinations would allow us to make a decision on which inducible promoter to use for Phase 3.
|2.2 Characterisation of light inducible promoters and RBS |
Aim: Varying combinations of light inducible promoters and RBS will be characterized to identify an efficient combination giving high levels of protein expression. In addition, constitutive promoters previously characterised will be used to express the light receptor protein YtvA required for efficient light sensing. In total, 4 combinations will be tried (Promoter-RBS),
|3.1 Characterisation of the clutch |
To characterise the effect of expression of the clutch, EpsE, on motility, EpsE is placed downstream of an inducible promoter.
3.2 Characterisation of Biomaterials
|Combinations of signal peptides (SP) and Biomaterials will be placed downstream of an inducible promoter to characterise the synthesis and secretion in B.subtilis. The following combinations were used (SP-Biomaterial):
|4.1 Light induced Expression of the clutch |
Aim: To test the blue light induced expression of the clutch, EpsE and the affct of this on motility.
|4.2 Light induced Expression of Biomaterials |
Aim: To test the blue light induced expression and secretion of Biomaterial.