BBa_B0012
1
BBa_B0012
TE from coliphageT7
2003-01-31T12:00:00Z
2015-08-31T04:07:20Z
Derived from the TE terminator of T7 bacteriophage between Genes 1.3 and 1.4 <genbank>V01146</genbank>.
Released HQ 2013
Transcription terminator for the <i>E.coli</i> RNA polymerase.
false
false
_1_
0
24
7
In stock
false
<P> <P>Suggested by Sri Kosuri and Drew Endy as a high efficiency terminator. The 5' end cutoff was placed immediately after the TAA stop codon and the 3' end cutoff was placed just prior to the RBS of Gene 1.4 (before AAGGAG).<P> Use anywhere transcription should be stopped when the gene of interest is upstream of this terminator.
false
Reshma Shetty
annotation1690
1
polya
range1690
1
28
41
annotation1686
1
T7 TE
range1686
1
8
27
annotation1687
1
stop
range1687
1
34
34
annotation7020
1
BBa_B0012
range7020
1
1
41
BBa_K142041
1
BBa_K142041
arabinose controlled RMF generator
2008-10-29T12:00:00Z
2015-05-08T01:10:22Z
Entire part was synthesized by Geneart.
This BioBrick expresses ribosome modulation factor (for details on RMF see either [http://partsregistry.org/wiki/index.php?title=Part:BBa_K142040 K142040] or the [http://2008.igem.org/Team:ETH_Zurich/Wetlab/Switch_Circuit ETH iGEM 2008 Wiki]upon induction with arabinose.
false
false
_194_
0
3254
9
Not in stock
false
No optimization was necessary.
false
Julius Rabl
component1995365
1
BBa_K142040
component1995370
1
BBa_B0011
component1995360
1
BBa_R0080
component1995366
1
BBa_B0012
component1995362
1
BBa_B0030
annotation1995370
1
BBa_B0011
range1995370
1
407
452
annotation1995362
1
BBa_B0030
range1995362
1
158
172
annotation1995360
1
BBa_R0080
range1995360
1
1
149
annotation1995365
1
BBa_K142040
range1995365
1
179
349
annotation1995366
1
BBa_B0012
range1995366
1
358
398
BBa_K142040
1
BBa_K142040
ribosome modulation factor (RMF)
2008-10-15T11:00:00Z
2015-05-08T01:10:22Z
The nucleotide sequence of RMF from Escherichia coli MG1655 was obtained from GenBank and used for primer design. Amplification from DH5-alpha genomic DNA template and addition of BioBrick standard prefix and suffix RMF was performed by PCR using the following primers:
forward primer
5'-cgcggaattcgcggccgcttctagatgaagagacaaaaacgagatcgcctgg
reverse primer
5'-cgcgctgcagcggccgctactagtattattaggccattactaccctgtccgc
The PCR product was subcloned into vector pSB1A7.
Short description: RMF is a 6.5kDa protein from E. coli, which upon expression rapidly terminates protein translation by binding to the ribosome active center. RMF can be removed and degraded by the cell, making its action reversible. Expression of RMF can be utilized to stop protein production in the cell if cell growth or the transcriptional responses of the cell are actively interfering with any planned genetic circuit. It enables the genetic engineer to turn the cell from an actively responding biological network into a mere test tube for any chosen duration of time. It might be of interest for certain applications that the amount of RMF molecules in the cell is unlikely to exceed the number of ribosomes.
Detailed description: Bacteria like E. coli terminate protein synthesis rapidly upon reaching the stationary phase of growth. Termination of translation is associated with dimerization of bacterial ribosomes; dimers (sedimentation velocity 100S as compared to 70S for standard monomeric ribosome) can be separated from monomers by sucrose gradient ultracentrifugation in low salt buffer, but dissociate during negative staining for transmission electron microscopy unless considerable amounts of crosslinking agents are used. Analysis of stationary phase ribosomes in E. coli has shown that several factors bind to the ribosome upon dimerization including RMF (1) and the hibernation promoting factor (2). RMF is expressed under a growth-phase dependent promotor (3) and sufficient to induce ribosome dimerization and termination of translation (4). Consequently, cells lacking RMF are unable to form dimers and have a markedly reduced survival rate in the stationary phase (5, 6). The structure of RMF has been determined by nuclear magnetic resonance (7) and biochemical crosslinking experiments have identified the binding site on the ribosome close to the peptidyl transferase center (8). Overexpression of RMF rapidly terminates protein production and growth of the cell; however unlike toxic proteins it does not kill the cell. Since RMF is a native constituent of the ribosome hibernation pathway it is assumed to be the least harmful way for turning off cell growth and protein production rapidly if cell viability is important for subsequent experiments.
References
(1) Wada, A., Yamazaki, Y., Fujita, N., and Ishihama, A. (1990) Structure and probable genetic location of a "ribosome modulation factor" associated with 100S ribosomes in stationary-phase Escherichia coli cells. Proc Natl Acad Sci U S A 87, 2657-61.
(2) Ueta, M., Ohniwa, R. L., Yoshida, H., Maki, Y., Wada, C., and Wada, A. (2008) Role of HPF (hibernation promoting factor) in translational activity in Escherichia coli. J Biochem 143, 425-33.
(3) Yamagishi, M., Matsushima, H., Wada, A., Sakagami, M., Fujita, N., and Ishihama, A. (1993) Regulation of the Escherichia coli rmf gene encoding the ribosome modulation factor: growth phase- and growth rate-dependent control. Embo J 12, 625-30.
(4) Wada, A., Igarashi, K., Yoshimura, S., Aimoto, S., and Ishihama, A. (1995) Ribosome modulation factor: stationary growth phase-specific inhibitor of ribosome functions from Escherichia coli. Biochem Biophys Res Commun 214, 410-7.
(5) El-Sharoud, W. M., and Niven, G. W. (2007) The influence of ribosome modulation factor on the survival of stationary-phase Escherichia coli during acid stress. Microbiology 153, 247-53.
(6) Yoshida, H., Maki, Y., Kato, H., Fujisawa, H., Izutsu, K., Wada, C., and Wada, A. (2002) The ribosome modulation factor (RMF) binding site on the 100S ribosome of Escherichia coli. J Biochem 132, 983-9.
(7) Tang, Y., Rossi, P., Swapna, G., Wang, H., Jiang, M., Cunningham, K., Owens, L., Ma, L., Xiao, R., Liu, J., Baran, M.C., Acton, T.B., Rost, B., Montelione, G.T. . (2007) Solution NMR Structure of Ribosome Modulation Factor VP1593 from Vibrio parahaemolyticus. Unpublished work.
(8) Yoshida, H., Yamamoto, H., Uchiumi, T., and Wada, A. (2004) RMF inactivates ribosomes by covering the peptidyl transferase centre and entrance of peptide exit tunnel. Genes Cells 9, 271-8.
false
false
_194_
0
3254
9
Not in stock
false
RMF was amplified from E. coli, therefore sequence optimization was unnecessary. The RMF gene does not contain any restriction sites for EcoRI, XbaI, SpeI or PstI and could be turned into a BioBrick without further modification.
false
Julius Rabl
annotation1981767
1
ribosome modulation factor
range1981767
1
1
171
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.
annotation7025
1
BBa_B0030
range7025
1
1
15
annotation1702
1
RBS
range1702
1
8
12
annotation1701
1
RBS-1\Strong
range1701
1
1
15
BBa_B0011
1
BBa_B0011
LuxICDABEG (+/-)
2003-01-31T12:00:00Z
2015-08-31T04:07:20Z
Derived from luxICDABEG operon terminator of Vibrio fischeri <genbank>AF170104</genbank>.
Released HQ 2013
Bidirectional transcriptional terminator consisting of a 22 bp stem-loop.</p>
false
false
_1_
0
24
7
In stock
false
<P> <P>In the naturally-occuring sequence there is a mismatch in the stem of the stem loop. This can be corrected via an A->G mutation (at position 40 -- sequence coordinate/not MFOLD coordinate). The above sequence does not reflect this mutation (but the MFOLD image does). This terminator's location cannot be found using some inverted repeat detectors like PALINDROME because it is too short and contains a mismatch. This one was found with the help of Tom Knight. It lies between two coding regions that point towards eachother.<P>
true
Reshma Shetty
annotation7019
1
BBa_B0011
range7019
1
1
46
annotation1683
1
stem_loop
range1683
1
13
35
BBa_R0080
1
AraC
Promoter (AraC regulated)
2004-01-27T12:00:00Z
2015-05-08T01:14:15Z
GenBank: J01641 (www.ncbi.nlm.nih.gov)
Released HQ 2013
AraC operator, truncated to include araO1, araI1, araI2, c-amp1, and c-amp2 sites. This operator should *activate* transcription in the presence of AraC; b/c the operator lacks the araO2 site, there should not be araC-mediated repression.
false
false
_1_
0
24
7
In stock
false
true
Sara Neves (Fighting Darwins)
annotation301456
1
c-amp2
range301456
1
4
29
annotation301457
1
araO1
range301457
1
6
44
annotation301462
1
ara1 and ara2
range301462
1
73
101
annotation308602
1
-10
range308602
1
136
141
annotation308601
1
-35
range308601
1
113
118
annotation301458
1
c-amp1
range301458
1
43
72
BBa_K142040_sequence
1
atgaagagacaaaaacgagatcgcctggaacgggcacatcaacgtggttatcaggccggcatcgccggacgctcaaaagaaatgtgtccctatcagacgctgaatcaaaggtcacaatggctgggaggctggcgagaagccatggcggacagggtagtaatggcctaataa
BBa_K142041_sequence
1
gcgtaacaaaagtgtctataatcacggcagaaaagtccacattgattatttgcacggcgtcacactttgctatgccatagcatttttatccataagattagcggatcctacctgacgctttttatcgcaactctctactgtttctccattactagagattaaagaggagaaatactagatgaagagacaaaaacgagatcgcctggaacgggcacatcaacgtggttatcaggccggcatcgccggacgctcaaaagaaatgtgtccctatcagacgctgaatcaaaggtcacaatggctgggaggctggcgagaagccatggcggacagggtagtaatggcctaataatactagagtcacactggctcaccttcgggtgggcctttctgcgtttatatactagagagagaatataaaaagccagattattaatccggcttttttattattt
BBa_B0030_sequence
1
attaaagaggagaaa
BBa_B0011_sequence
1
agagaatataaaaagccagattattaatccggcttttttattattt
BBa_B0012_sequence
1
tcacactggctcaccttcgggtgggcctttctgcgtttata
BBa_R0080_sequence
1
gcgtaacaaaagtgtctataatcacggcagaaaagtccacattgattatttgcacggcgtcacactttgctatgccatagcatttttatccataagattagcggatcctacctgacgctttttatcgcaactctctactgtttctccat
igem2sbol
1
iGEM to SBOL conversion
Conversion of the iGEM parts registry to SBOL2.1
James Alastair McLaughlin
Chris J. Myers
2017-03-06T15:00:00.000Z