BBa_K1783002
1
BBa_K1783002
Constitutive Promoter-RBS-Unstable RFP (LVA-Tagged)
2015-07-17T11:00:00Z
2015-09-25T10:32:37Z
Promoter, RBS, LVA tag from E. coli
RFP from Discosoma striata
Produces unstable RFP under a constitutive promoter.
false
false
_2208_
26311
20382
9
false
none
false
Chun Mun Loke, Adam Wahab, Kimia Abtahi, Iowis Zhu
component2433285
1
BBa_K081005
component2433291
1
BBa_K1399001
annotation2433291
1
BBa_K1399001
range2433291
1
65
778
annotation2433285
1
BBa_K081005
range2433285
1
1
58
BBa_K1399001
1
BBa_K1399001
RFP from Discosoma striata (coral) with LVA-ssrA degradation tag
2014-09-17T11:00:00Z
2015-05-08T01:10:15Z
RFP comes from part BBa_E1010, tag sequence was obtained from paper by Andersen et al., (1998).[2]
Mutant RFP from Discosoma striata (coral) (see part BBa_E1010) with added LVA-ssrA degradation tag. The tag increases RFP turn-over rate, thus providing better temporal resolution of red fluorescence. In the same time, maximal fluorescence amplitudes will be lower as newly formed protein is degraded as soon as it is formed.
This tag is commonly attached to repressor proteins for use in various gene networks (e.g., oscillators).
The tag encodes peptide sequence AANDENYALVA and is recognized by ClpA and ClpX unfoldases and ClpX mediator SspB.[1] ClpA and ClpX then form a proteosome-like complex with ClpP protease and the protein is degraded.[1]
The final three residues of the tag determines the strength of interaction with ClpX and thus the final protein degradation rate.[2] The LVA tag is reported to lead to fast protein degradation, degrading GFP with rate -0.018 per minute.[2] However, be aware that exact protein degradation rate depends on multiple factors: ClpXP and ClpAP protease and SspB mediator concentrations; protein stability; Km of binding to the protease; temperature [3].
References:
[1] Flynn, J. M. et al. Overlapping recognition determinants within the ssrA degradation tag allow modulation of proteolysis. Proc. Natl. Acad. Sci. U. S. A. 98, 10584???9 (2001).
[2] Andersen, J. B. et al. New unstable variants of green fluorescent protein for studies of transient gene expression in bacteria. Appl. Environ. Microbiol. 64, 2240???6 (1998).
[3] Purcell, O., Grierson, C. S., Bernardo, M. Di & Savery, N. J. Temperature dependence of ssrA-tag mediated protein degradation. J. Biol. Eng. 6, 10 (2012).
false
false
_1777_
0
22477
9
In stock
true
The tag was attached to RFP using PCR and MABEL (mutagenesis with blunt-end ligation), thus avoiding introduction of additonal residues and restriction sites. Different parts of the tag are recognized by different proteins, for example, the final 3 residues (LVA in this case) are recognised by ClpX, whereas first 4 residues of the tag are required for efficient SspB binding.[1] Thus modifications of these critical residues alter the efficacy with what different proteases bind to it.
false
Anna Stikane
annotation2383875
1
stop
range2383875
1
709
711
annotation2383872
1
start
range2383872
1
1
3
annotation2383876
1
stop
range2383876
1
712
714
annotation2383873
1
cds
range2383873
1
4
675
annotation2383874
1
LVA-ssrA tag
range2383874
1
676
708
BBa_K081005
1
BBa_K081005
constitutive promoter family member and RBS
2008-10-17T11:00:00Z
2015-05-08T01:08:34Z
Promoter: John Anderson.
RBS: Vinay S Mahajan, Voichita D. Marinescu, Brian Chow, Alexander D Wissner-Gross and Peter Carr IAP, 2003.
Released HQ 2013
Constitutive promoter (strong) with RBS (strong, efficiency=0.3)
false
true
_227_
0
2583
9
In stock
true
We used BioBrick Standard Assembly.
true
Lorenzo Pasotti, Paolo Magni
component1981863
1
BBa_J23100
component1981865
1
BBa_B0030
annotation1981865
1
BBa_B0030
range1981865
1
44
58
annotation1981863
1
BBa_J23100
range1981863
1
1
35
BBa_J23100
1
BBa_J23100
constitutive promoter family member
2006-08-03T11:00:00Z
2015-08-31T04:08:40Z
Isolated from library of promoters
Released HQ 2013
Replace later
false
true
_52_
0
483
95
In stock
true
N/A
true
John Anderson
BBa_B0030
1
BBa_B0030
RBS.1 (strong) -- modified from R. Weiss
2003-01-31T12:00:00Z
2015-08-31T04:07:20Z
Released HQ 2013
Strong RBS based on Ron Weiss thesis. Strength is considered relative to <bb_part>BBa_B0031</bb_part>, <bb_part>BBa_B0032</bb_part>, <bb_part>BBa_B0033</bb_part>.
false
true
_44_46_
0
24
7
In stock
false
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix ("orig" in figure 4-14 of Ron Weiss thesis). <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 <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.
annotation1702
1
RBS
range1702
1
8
12
annotation1701
1
RBS-1\Strong
range1701
1
1
15
annotation7025
1
BBa_B0030
range7025
1
1
15
BBa_J23100_sequence
1
ttgacggctagctcagtcctaggtacagtgctagc
BBa_K1399001_sequence
1
atggcttcctccgaagacgttatcaaagagttcatgcgtttcaaagttcgtatggaaggttccgttaacggtcacgagttcgaaatcgaaggtgaaggtgaaggtcgtccgtacgaaggtacccagaccgctaaactgaaagttaccaaaggtggtccgctgccgttcgcttgggacatcctgtccccgcagttccagtacggttccaaagcttacgttaaacacccggctgacatcccggactacctgaaactgtccttcccggaaggtttcaaatgggaacgtgttatgaacttcgaagacggtggtgttgttaccgttacccaggactcctccctgcaagacggtgagttcatctacaaagttaaactgcgtggtaccaacttcccgtccgacggtccggttatgcagaaaaaaaccatgggttgggaagcttccaccgaacgtatgtacccggaagacggtgctctgaaaggtgaaatcaaaatgcgtctgaaactgaaagacggtggtcactacgacgctgaagttaaaaccacctacatggctaaaaaaccggttcagctgccgggtgcttacaaaaccgacatcaaactggacatcacctcccacaacgaagactacaccatcgttgaacagtacgaacgtgctgaaggtcgtcactccaccggtgctgctgcaaacgacgaaaactacgctttagtagcttaataa
BBa_K081005_sequence
1
ttgacggctagctcagtcctaggtacagtgctagctactagagattaaagaggagaaa
BBa_B0030_sequence
1
attaaagaggagaaa
BBa_K1783002_sequence
1
ttgacggctagctcagtcctaggtacagtgctagctactagagattaaagaggagaaatactagatggcttcctccgaagacgttatcaaagagttcatgcgtttcaaagttcgtatggaaggttccgttaacggtcacgagttcgaaatcgaaggtgaaggtgaaggtcgtccgtacgaaggtacccagaccgctaaactgaaagttaccaaaggtggtccgctgccgttcgcttgggacatcctgtccccgcagttccagtacggttccaaagcttacgttaaacacccggctgacatcccggactacctgaaactgtccttcccggaaggtttcaaatgggaacgtgttatgaacttcgaagacggtggtgttgttaccgttacccaggactcctccctgcaagacggtgagttcatctacaaagttaaactgcgtggtaccaacttcccgtccgacggtccggttatgcagaaaaaaaccatgggttgggaagcttccaccgaacgtatgtacccggaagacggtgctctgaaaggtgaaatcaaaatgcgtctgaaactgaaagacggtggtcactacgacgctgaagttaaaaccacctacatggctaaaaaaccggttcagctgccgggtgcttacaaaaccgacatcaaactggacatcacctcccacaacgaagactacaccatcgttgaacagtacgaacgtgctgaaggtcgtcactccaccggtgctgctgcaaacgacgaaaactacgctttagtagcttaataa
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