BBa_K1991007
1
BBa_K1991007
Pcons-RBS-LO-BamHI
2016-10-11T11:00:00Z
2016-10-12T03:28:23Z
E. coli (Lpp-OmpA)
A study showed that Lpp-OmpA (LO) hybrid can direct the heterologous protein GFP to the external surface of E. coli (Enzyme Microb Technol. 2001). Bacillus lipase (J Microbiol. 2014) and Fungi xylanase (Curr Microbiol. 2015) were demonstrated to be displayed on the cell surface of E. coli and maintained the functional enzyme activities. We???d like to display the AOX enzyme on bacterial surface by fusing with Lpp-OmpA and apply to the electrochemical analyzer by depositing on the test strip.
false
false
_2458_
31417
31417
9
false
No
false
Chen, Pei-En
component2493570
1
BBa_K1991004
component2493567
1
BBa_J23101
component2493569
1
BBa_B0034
annotation2493570
1
BBa_K1991004
range2493570
1
62
502
annotation2493567
1
BBa_J23101
range2493567
1
1
35
annotation2493569
1
BBa_B0034
range2493569
1
44
55
BBa_K1991004
1
BBa_K1991004
Lpp-OmpA-BamHI
2016-10-09T11:00:00Z
2016-10-12T03:03:40Z
Modified based on BBa_K1694002 (Lpp-OmpA-N)
Lpp and OmpA are outer membrane proteins of E. coli. Lpp-OmpA (LO) hybrid can direct heterologous proteins to bacterial cell surface. In 2015, NCTU-Formosa used it to display scFv (single chain fragment variable) antibodies on the surface of E. coli. They found that a fusion protein cannot be possible to be created under the standard BioBrick assembly rule, that is EcoRI(E)-XbaI(X)-GENE-SpeI(S)-PstI(P). The A part of EX-LO-SP and the B part of EX-scFV-SP, for example, are connected by cutting and ligation of SpeI plus PstI for the A part and XbaI plus PstI for the B part. The SCAR generated by XbaI/SpeI (ACTAGA) will form a stop codon just in front of the ATG start codon of the scFV protein of the B part. This situation has been officially mentioned by the BioBrick standard assembly.
Therefore, in 2015, NCTU-Formosa created a novel part (BBa_K1694002) putting NcoI site between the LO part and SpeI site. However, when considering cloning, we found that an extra NcoI site is present on Cm resistance gene making it difficult be a vector for gene cloning.
In 2016, Mingdao improved the part by replacing NcoI site with BamHI site (BBa_K1991004). Also, we???ve confirmed and prove the function of LO directing a fusion protein to the cell surface with enzyme activity in our project.
false
false
_2458_
31417
31417
9
false
BBa_K1991004: Lpp-OmpA-BamHI/pSB1C3
The DNA fragment of Lpp-OmpA was amplified by PCR using BBa_K1694035 as a template with a primer containing BamHI site and digested by EcoRI and SpeI, followed by cloning onto pSB1C3 which was cut by EcoRI and SpeI. The part has been confirmed by sequencing.
false
Chen, Pei-En
annotation2521542
1
Lpp
range2521542
1
1
87
annotation2521543
1
linker
range2521543
1
88
90
annotation2521545
1
BamHI
range2521545
1
436
441
annotation2521544
1
OmpA
range2521544
1
91
435
BBa_J23101
1
BBa_J23101
constitutive promoter family member
2006-08-03T11:00:00Z
2015-08-31T04:08:40Z
later
Released HQ 2013
later
false
true
_52_
0
483
95
In stock
true
N/A
true
John Anderson
BBa_B0034
1
BBa_B0034
RBS (Elowitz 1999) -- defines RBS efficiency
2003-01-31T12:00:00Z
2015-08-31T04:07:20Z
Released HQ 2013
RBS based on Elowitz repressilator.
false
true
_1_
0
24
7
In stock
false
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix. <p>No secondary structures are formed in the given RBS region. Users should check for secondary structures induced in the RBS by upstream and downstream elements in the +50 to -50 region, as such structures will greatly affect the strength of the RBS.
Contact info for this part: <a href="mailto:(bchow@media.mit.edu)">Brian Chow</a>
true
Vinay S Mahajan, Voichita D. Marinescu, Brian Chow, Alexander D Wissner-Gross and Peter Carr IAP, 2003.
annotation23325
1
conserved
range23325
1
5
8
BBa_B0034_sequence
1
aaagaggagaaa
BBa_K1991004_sequence
1
atgaaagcaaccaagctggttctgggtgccgtgattctgggcagtaccctgttagcaggttgttctagcaatgccaaaatcgaccaaggcatcaacaacaatggcccgacccacgaaaaccagctgggtgccggtgcctttggtggttatcaggtgaacccgtacgtgggctttgaaatgggctatgattggctgggccgcatgccgtacaaaggcagtgtggagaacggcgcctataaagcacagggcgtgcagctgacagcaaaactgggctaccctattaccgacgacctggacatctacacacgcttaggcggcatggtgtggcgcgccgataccaagagcaacgtgtacggcaagaaccacgataccggcgtgagtccggtgtttgccggcggtgtggagtatgcaatcaccccggaaattgccacacgtggatcc
BBa_J23101_sequence
1
tttacagctagctcagtcctaggtattatgctagc
BBa_K1991007_sequence
1
tttacagctagctcagtcctaggtattatgctagctactagagaaagaggagaaatactagatgaaagcaaccaagctggttctgggtgccgtgattctgggcagtaccctgttagcaggttgttctagcaatgccaaaatcgaccaaggcatcaacaacaatggcccgacccacgaaaaccagctgggtgccggtgcctttggtggttatcaggtgaacccgtacgtgggctttgaaatgggctatgattggctgggccgcatgccgtacaaaggcagtgtggagaacggcgcctataaagcacagggcgtgcagctgacagcaaaactgggctaccctattaccgacgacctggacatctacacacgcttaggcggcatggtgtggcgcgccgataccaagagcaacgtgtacggcaagaaccacgataccggcgtgagtccggtgtttgccggcggtgtggagtatgcaatcaccccggaaattgccacacgtggatcc
igem2sbol
1
iGEM to SBOL conversion
Conversion of the iGEM parts registry to SBOL2.1
Chris J. Myers
James Alastair McLaughlin
2017-03-06T15:00:00.000Z