Difference between revisions of "Part:BBa K103006"
(→results) |
(→Usage and Biology) |
||
| Line 13: | Line 13: | ||
*In our project Ompa-link is used as outer membrane anchor for our selection system | *In our project Ompa-link is used as outer membrane anchor for our selection system | ||
*This brick contains our nonstandard restriction sites (NdeI and SacI) that allow 'scarless' cloning and easy creation of translation fusions | *This brick contains our nonstandard restriction sites (NdeI and SacI) that allow 'scarless' cloning and easy creation of translation fusions | ||
| + | | [[Image:Ompalinker.jpg|200px]] | ||
| + | This protein structure was predicted using threading and protein fold prediction and may differ from actual one. | ||
| + | |} | ||
| + | |||
===improvement by NEFU-China=== | ===improvement by NEFU-China=== | ||
Lpp-ompA, a signal peptide can be used as a outer-membrane-targeting anchor in E.coli. Our team modified the signal peptide sequence to make it more conducive to direct our FABP to anchor to the outer membrane of E.coli. We reduced several amino acids at the N-end of the peptide chain so that the sequence was shorter than that in the Registry of iGEM, BBa_K103006, from University of Warsaw 2008 iGEM team, but the FABP with our signal peptide was expressed in high level. In the meantime, we used codon optimization to further improve the expressed level of FABP. The result showed that expressed levels of FABP with our shorter signal peptides were higher than BBa_K103006 in the Registry of iGEM. | Lpp-ompA, a signal peptide can be used as a outer-membrane-targeting anchor in E.coli. Our team modified the signal peptide sequence to make it more conducive to direct our FABP to anchor to the outer membrane of E.coli. We reduced several amino acids at the N-end of the peptide chain so that the sequence was shorter than that in the Registry of iGEM, BBa_K103006, from University of Warsaw 2008 iGEM team, but the FABP with our signal peptide was expressed in high level. In the meantime, we used codon optimization to further improve the expressed level of FABP. The result showed that expressed levels of FABP with our shorter signal peptides were higher than BBa_K103006 in the Registry of iGEM. | ||
Revision as of 09:05, 27 October 2017
OmpA outer membrane protein A fused to linker; displays proteins on cell surface
One of our basic bricks used to create fusions attached to outer membrane
Usage and Biology
|
This protein structure was predicted using threading and protein fold prediction and may differ from actual one. |
improvement by NEFU-China
Lpp-ompA, a signal peptide can be used as a outer-membrane-targeting anchor in E.coli. Our team modified the signal peptide sequence to make it more conducive to direct our FABP to anchor to the outer membrane of E.coli. We reduced several amino acids at the N-end of the peptide chain so that the sequence was shorter than that in the Registry of iGEM, BBa_K103006, from University of Warsaw 2008 iGEM team, but the FABP with our signal peptide was expressed in high level. In the meantime, we used codon optimization to further improve the expressed level of FABP. The result showed that expressed levels of FABP with our shorter signal peptides were higher than BBa_K103006 in the Registry of iGEM.
results
E.coli was induced in 37℃ and membrane protein was separated by a speeding centrifuge. The result of western blot showed the difference at the expressed levels of Lpp-ompA-L-FABP in E.coli transformed by our recombinant plasmid and plasmid in registry of iGEM.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]