Part:BBa_K5387002:Design

Design Notes

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.