Difference between revisions of "Part:BBa K3484006"
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=Characterization experiments= | =Characterization experiments= | ||
− | For the characterization | + | For the characterization a Plate-Reader analysis was made. All the information on the experimental conditions and parameters used are described on the table below (Tab.9). |
Revision as of 23:15, 25 October 2020
sfGFP + ASV tag
The superfolder green fluorescent protein (sfGFP) is a variant of the common green fluorescent protein, that folds faster, thus generating fluorescent in less time.
Model Characterization
In order to quantify how the ASV tag affected the degradation rate of the sfGFP, a simple mathematical model was developed. The following model is exactly the same for the two scenarios, and consists of a constitutive expression minus a degradation rate (Equation.1). As both systems are constitutively regulated by the same promoter, the PJ23100 (BBa_J23100), the production rate will be the same in the two systems. In addition, the equations representing the steady-state (dy/dx=0) of the system were calculated (Equation.2).
With the Steady-State equations we are able to look for the relationship between the two degradation rates (δ) as the production rate is the same(Equation.3).
Characterization experiments
For the characterization a Plate-Reader analysis was made. All the information on the experimental conditions and parameters used are described on the table below (Tab.9).
Table 1. Plate-Reader Parameters for the characterization of the effects of ASV tag in sfGFP | |||
Parameters | Value | ||
Plate-Reader model | Synergy HTX | ||
Plate type | Thermo Fischer 96-well microplates black-walled clear bottom | ||
Cell medium | LB | ||
Time | 24 hours | ||
Shake | Linear: Continuous, Frequency: 567 rpm (3mm) | ||
Temperature | 37C | ||
Gain | 50 | ||
Optical Density (OD) measurement (absorbance) | 660nm | ||
GFP excitation wavelength | 485nm | ||
GFP emission wavelength | 528nm |
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 25
Illegal AgeI site found at 148 - 1000COMPATIBLE WITH RFC[1000]