Difference between revisions of "Part:BBa K5387002:Design"
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===Design Notes=== | ===Design Notes=== | ||
| + | The gene has been oprimized for expression in ''E. coli''. | ||
| + | |||
Some parts of the protein are membrane bound, which makes them much harder to solve and express in ''E. coli''. We therefore tried various different primers to try to clone different parts of the protein to pET28a(+), listed below: | Some parts of the protein are membrane bound, which makes them much harder to solve and express in ''E. coli''. We therefore tried various different primers to try to clone different parts of the protein to pET28a(+), listed below: | ||
Revision as of 12:49, 20 August 2024
Design Notes
The gene has been oprimized for expression in E. coli.
Some parts of the protein are membrane bound, which makes them much harder to solve and express in E. coli. We therefore tried various different primers to try to clone different parts of the protein to pET28a(+), listed below:
To clone the whole sequence into pET28a(+) between the NdeI and EcoRI restriction sites:
- fwd: 5'-GCGCCATATGATGGACGGACCCAGGTCAG-3'
- rev: 5'-GCGCGAATTCTTAGGCGCCACCTCTGCTTGCCATC-3'
To delete the first 100 amino acids:
- fwd: 5'-GCGCCATATGGGTATGCTTGTGGTCAACGG-3'
- rev: 5'-GCGCGAATTCTTAGGCGCCACCTCTGCTTGCCATC-3'
To delete the last 30 amino acids:
- fwd: 5'-GCGCCATATGATGGACGGACCCAGGTCAG-3'
- rev: 5'-CAGGAGCCACGTCAACCTGAATAACACC-3'
To delete the first 100 and last 30 and amino acids:
- fwd: 5'-GCGCCATATGGGTATGCTTGTGGTCAACGG-3'
- rev: 5'-CAGGAGCCACGTCAACCTGAATAACACC-3'
Source
The genes is expressed in human cells.