Difference between revisions of "Part:BBa K3117026"

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===Usage and Biology===

===Usage and Biology===

The sequence contains a C-terminal His-tag (BBa_K3117005) for easy purification and detection. Secretion of the protein is ensured by an Igk leader (BBa_K3117006), which directs the produced protein into the secretory pathway. When the protein passes the membrane, the leader segment is cleaved off. By connecting the variable regions of the heavy and the light chain of an anti-GPA33 antibody with a short, flexible GGGGS linker (BBa_K3117028), the scFv retains its antigen-binding ability and is much smaller than a conventional antibody.

The sequence contains a C-terminal His-tag (BBa_K3117005) for easy purification and detection. Secretion of the protein is ensured by an Igk leader (BBa_K3117006), which directs the produced protein into the secretory pathway. When the protein passes the membrane, the leader segment is cleaved off. By connecting the variable regions of the heavy and the light chain of an anti-GPA33 antibody with a short, flexible GGGGS linker (BBa_K3117028), the scFv retains its antigen-binding ability and is much smaller than a conventional antibody.

Therefore, it was cloned into the expression vector pSEC/tag2/Hygro\C\-\OK by Gibson Assembly. Our protein was under control of the CMV promotor ensuring a high-level constitutive expression. Afterwards, the construct was brought into HEK293T cells with either calcium phosphate transfection or lipofection.

Therefore, it was cloned into the expression vector pSEC/tag2/Hygro\C\-\OK by Gibson Assembly. Our protein was under control of the CMV promotor ensuring a high-level constitutive expression. Afterwards, the construct was brought into HEK293T cells with either calcium phosphate transfection or lipofection.

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