Three complete c-di-GMP riboswitches (Bc3, Bc4 and Bc5 RNA) with similar structures, which are arranged in tandem to constitute a triple-tandem (Bc3-5 RNA) riboswitch in the 5′-UTR of the cspABCDE mRNA in Bacillus thuringiensis subsp. chinensis CT-43.
We designed the sequence of bc3-5 by add perfix and suffix which are belong to BioBrick RFC 10.
From Bacillus thuringiensis subsp. chinensis CT-43 In the CT-43 genome19,20 (GenBank accession: NC_017208.1), five type I c-di-GMP riboswitches named Bc1, Bc2, Bc3, Bc4 and Bc5 RNA (Supplementary Tables S1???S5) were annotated in the Rfam database (http://rfam.sanger.ac.uk/search). Zhou, H., et al. (2016). "Characterization of a natural triple-tandem c-di-GMP riboswitch and application of the riboswitch-based dual-fluorescence reporter." Sci Rep 6: 20871.
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene II message (BBa_M13002). The part aligns with base pairs 8252-8267 in M13K07 genome. It was identified as the RBS for gene II message by Wezenbeek et al in Gene (1980) 11: 129-148
New England Biolabs M13K07. Sequenced in 2002 by L. Panganaban
Variant of GFP that folds robustly even when fused to poorly folded proteins. Waldo and other research groups has reported the engineering of a superfolder GFP (sfGFP) that showed increased resistance to denaturation, improved folding kinetics, and increased resistance to aggregation during refolding (Pedelacq et al., 2006; Andrews et al., 2007; Fisher and DeLisa, 2008). sfGFP has proven to be very useful as a scaffold for improved protein detection and tagging both in vivo and in vitro using self-assembled sfGFP fragments (Cabantous et al., 2005b; Cabantous and Waldo, 2006). Furthermore, sfGFP fusions are more soluble than conventional GFP fusions (Wu et al., 2009).
Reverse RBS (RBSrev) -- corresponds to BBa_B0030
Repeats in oligos caused unusual products during cloning
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene VII message (BBa_M13007). The part aligns with base pairs 1092-1107 in M13K07 genome. It was identified as the RBS for gene VII message by Wezenbeek et al in Gene (1980) 11: 129-148
The ribosomal binding site of the creD gene in the E Coli creD operon.
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene III message (BBa_M13003). The part aligns with base pairs 1563-1578 in M13K07 genome. It was identified as the RBS for gene III message by Wezenbeek et al in Gene (1980) 11: 129-148 New England Biolabs M13K07. Sequenced in 2002 by L. Panganaban
Sequence comes from the natural gene 5 RBS. Using gene 5 as a strong RBS is from initial results from experiments characterizing all natural T7 RBSs.
RBS (Elowitz 1999) -- defines RBS efficiency
Vinay S Mahajan, Voichita D. Marinescu, Brian Chow, Alexander D Wissner-Gross and Peter Carr IAP, 2003.
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix. 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: Brian Chow
trkD, a functional Kup (formerly TrkD) system took up Cs+ with a moderate rate and affinity
Escherichia coli cells which contain a functional Kup (formerly TrkD) system took up Cs' with a moderate rate and affinity. Kup is a separate K+ uptake system with relatively little discrimination in the transport of the cations K+, Rb+, and Cs'. Regardless of the presence or absence of Kup, K+-replete cells took up Cs'primarily by a very low affinity mode, proportional to the ratio of the Cs' and K+ concentrations in the medium. In our project, trkD is used to collect Cs-137 when recN is activated by radiation and start the transcription of trkD,in order to absorb this kind of radioactive source like Cs-137
This part is just for upload test. It is meaningless
b0034 derived RBS to match spacing of J01122
RBS spacing modification to [RBS] such that the spacing between the shine-delgarno and the ORF is the same as it is in J01122.
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene XI message (BBa_M13011). The part aligns with base pairs 3900-3915 in M13K07 genome. It was NOT identified as the RBS for gene XI message by Wezenbeek et al in Gene (1980) 11: 129-148
ribosome binding site for creA gene in the E Coli cre operon
Modified version of B0031 from the Voigt Lab. Not currently available as an independent part (only in a tet inverter).
Complete sMMO of Methylococcus capsulatus
The part consists of the six subunits of methane monooxygenase (MMO). Gene order: mmoX, mmoY, mmoB, mmoZ, mmoD, mmoC The expression is under control of lac+pl. MMO converts methane to methanol.
Part is designed considering the RFC10 standard.
Sequence comes from the natural B0030. Using B0030 as a medium strength RBS is from initial results from experiments characterizing all natural T7 RBSs.
RBS.1 (strong) -- modified from R. Weiss
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>.
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix ("orig" in figure 4-14 of Ron Weiss thesis). 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 Brian Chow
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene X message (BBa_M13010). The part aligns with base pairs 480-495 in M13K07 genome. It was identified as the RBS for gene II message by Wezenbeek et al in Gene (1980) 11: 129-148
Luminesensor repressible SulA408 promoter
SulA promoter in 408 form.SulA promoter belongs to the SOS regulon family, and promotes in E.coli transcription of SulA protein. SulA408 promoter is reponsive to our optimized Luminesensor(BBa_K819005).
SulA promoter sequence is got from regulon.org.
This is the native ribosome binding site of ftsK, a DNA-translocating motor protein involved in cell division in Escherichia coli.
Virtual part to be added to coding sequence BBa_I742137.
Medium RBS based on Ron Weiss thesis. Strength considered relative to <bb_part>BBa_B0030</bb_part>, <bb_part>BBa_B0032</bb_part>, <bb_part>BBa_B0033</bb_part>.
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix ("RBS-1" in figure 4-14 of thesis). 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: Brian Cho
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene IV message (BBa_M13004). The part aligns with base pairs 4204-4219 in M13K07 genome. It was identified as the RBS for gene IV message by Wezenbeek et al in Gene (1980) 11: 129-148
Synthetic ribosome binding site added by Son of Babel procedure
This is a 4bp synthetic ribosome binding site which can be added by fusion PCR using a reverse primer and the Son of Babel cloning system.
The part can be added by fusion PCR using a Son of Babel reverse primer.
ribosome binding site for transcription initiation
Construct the JBA (J23119+B0034+NhaA), B0034 and NhaB with the vector pSB1C3. Two functional genes NhaA and NhaB are used to pump in protons and lower the pH of bacteria???s microenvironment. JBA and JBB are linked with pSB1C3 through 3A Assemble successfully. Thus, it can be an alkaline-resisting part.
This is a composite part including the J23119 promoter ecoded in part BBa_J23119 and the ribosome binding site B0034 ecoded in part BBa_B0034. The functional genes NhaA and NhaB are cloned from Escherichia coli str. K-12 substr. sequence found through Genbank http://www.ncbi.nlm.nih.gov/nuccore/NC_000913.3?report=genbank&from=17489&to=18655 http://www.ncbi.nlm.nih.gov/nuccore/NC_000913.3?report=genbank&from=1233176&to=1234717&strand=true
The sequence of promoter J23119 is found through the standard part BBa_J23119 http://parts.igem.org/Part:BBa_J23119 The sequence of ribosome binding site B0034 is found through the standard part BBa_B0034 http://parts.igem.org/Part:BBa_B0034 The sequence of gene NhaA is found through Genbank http://www.ncbi.nlm.nih.gov/nuccore/NC_000913.3?report=genbank&from=17489&to=18655 The sequence of gene NhaB is found through Genbank http://www.ncbi.nlm.nih.gov/nuccore/NC_000913.3?report=genbank&from=1233176&to=1234717&strand=true
Constantin Ahlmann-Eltze, Charlotte Bunne, Magdalena B??scher, Jan Gleixner, Max Horn, Anna Huhn, Nils Klughammer, Jakob Kreft, Elisabeth Sch??fer, Carolin Schmelas, Silvan Schmitz, Max Waldha
RFC10 compatible strong RBS derived from the T7 phage gene 10a (major capsid protein)[1]. When assembled to a coding part with the A of the start codon being part of the XbaI site, the RBS will be shifted one bp downstream compared to the native sequence. The sequence was successfully used by the iGEM team Heidelberg 2014 for the expression of many proteins in E.coli. 1. Olinss, P. & Rangwala, S. H. Derived from Bacteriophage T7 mRNA Acts ELS an Enhancer of Translation of the lac2 Gene in. 16973???16976 (1989).
The 18 bp including the XbaI that can be found upstream of the presumably important part of the RBS were included into the sequence just to make sure it works. However to fully comply with RFC10 a G was inserted behind the XbaI site.
synthesized as found in the T7 genome and several commercial expression plasmids.
Test RBS from T7 that may be used in E.coli
Weak1 RBS based on Ron Weiss thesis. Strength is considered relative to BBa_B0030, BBa_B0031, BBa_B0033.
This version of the T7 0.3 RBS is designed for construction via Heather Keller's alternative assembly plan using XbaI and SapI for RBS/CDS assembly without a mixed site. Contains the -20 to -1 region of gp0.4 from the wild-type T7 genome. Can also be used with Standard BioBricks assembly, although this will add a 7 amino acid fusion to the N-terminus of the resulting protein.
This RBS is designed to be compatible with Heather Keller's RBS assembly scheme for creating RBS/CDS junctions without a mixed site. For use in this scheme, this part should be cut with XbaI and SapI. It must be used with a coding sequence containing the SapI site at the 5' end. This part is not compatible with CDSs that use a GTG site and must be used with CDSs using an ATG start site. The two bp spacer on the 3' end also allows this part to be used in standard biobricks assembly and is necessary to maintain the reading frame of a downstream CDS. Note that Standard Biobricks Assembly with this part results in the creation of a new ATG start site, and a 7 amino acid fusion at the N terminus of the downstream protein. CDSs with alternative start sites are compatible with this part via Standard Assembly. This part was inadvertently constructed without the single base pair (G/C) that separates the XbaI site and the 5' end of the part.
The RBS is taken from the wild-type bacteriophage T7 genome. Please see part B2001 for more information.
This is the -20 to -1 region of T7 gp0.5, which contains the native RBS region for this gene. The physical instantiation of this part does not exist. This part is not compatible with Standard Biobricks assembly methods, as the scar created during assembly would interrupt the spacing between the RBS and the start codon of the CDS. Individuals wishing to use this RBS sequence in their assembles should refer to part B2103.
The -20 to -1 region was chosen to be representative of the RBS region of the T7 genes for the following reasons: 1) The Shine-Delgarno region is contained within this defined region for all T7 genes. 2) Current literature suggests that when bound to the Translation Start Site (0), the Ribosome covers positions -20 to +13 of the gene.
Genome of wild-type bacteriophage T7, positions 1476-1495. Includes the -20 to -1 region of gp0.5.
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene VIII message (BBa_M13008). The part aligns with base pairs 1285-1300 in M13K07 genome. It was identified as the RBS for gene VIII message by Wezenbeek et al in Gene (1980) 11: 129-148
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene I message (BBa_M13001). The part aligns with base pairs 8252-8267 in M13K07 genome. It was identified as the RBS for gene II message by Wezenbeek et al in Gene (1980) 11: 129-148
strong synthetic E. coli ribosome binding site with SacI site.
This is a strong synthetic E. coli ribosome binding site. It is synthesised as two complementary oligonucleotides rather than being incorporated into a biobrick plasmid. It incorporates a SacI site overlapping the XbaI site, which is preserved when it is added to any other biobrick. This allows easy detection of the RBS after it has been added upstream of a biobrick coding sequence in a plasmid vector.
Note the presence of a SacI site overlapping the XbaI site, which is preserved when this biobrick is added to any other biobrick. At the time of writing, this biobrick is added as a short piece of DNA composed of two complementary oligonucleotides rather than being incorporated into a biobrick cloning vector by itself. It can be added upstream of a biobrick coding sequence, and its presence can easily be detected in miniprep DNA on a gel by using a SacI-SpeI or similar digest.
Strong RBS based on Ron Weiss thesis. Strength is considered relative to BBa_B0031, BBa_B0032, BBa_B0033 and BBa_B0034.
Biobricked version of Isaacs' riboregulator cis repressed lock, crR12.
This is the -20 to -1 region of T7 gp0.7, which contains the native RBS region for this gene. The physical instantiation of this part does not exist. This part is not compatible with Standard Biobricks assembly methods, as the scar created during assembly would interrupt the spacing between the RBS and the start codon of the CDS. Individuals wishing to use this RBS sequence in their assembles should refer to part XXXXX. (Information forthcoming)
The -20 to -1 region was chosen to be representative of the RBS region of the T7 genes for the following reasons: 1) The Shine-Delgarno region is contained within this defined region for all T7 genes. 2) Current literature suggests that when bound to the Translation Start Site (0), the Ribosome covers positions -20 to +13.
Genome of wild-type bacteriophage T7, positions 2001-2020. Includes the -20 to -1 region of gp0.7.
Ribosome binding site for transcription initiation
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene IX message (BBa_M13009). The part aligns with base pairs 1190-1205 in M13K07 genome. It was identified as the RBS for gene IX message by Wezenbeek et al in Gene (1980) 11: 129-148
This is a single bp change from B0034. It's strength is approximately 35% that of B0034.
Restriction sites of Sal I, Bam HI, Bgl II, and Mlu I in front of strong RBS
We designed new prefix(Sal I and Bam HI) and suffix(Bgl II and Mul I) for insertions of promoters and regulator binding sites in front of RBS.
The ribosomal binding site for gene creC in the E Coli cre operon.
RBS.3 (medium) -- derivative of BBa_0030
Weak1 RBS based on Ron Weiss thesis. Strength is considered relative to <bb_part>BBa_B0030</bb_part>, <bb_part>BBa_B0031</bb_part>, <bb_part>BBa_B0033</bb_part>.
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix ("RBS-2" in figure 4-14 of thesis). Contact info for this part: Brian Chow
Annexin V is member of the annexin family of Ca dependent proteins. Its function is not yet known. It binds to phosphatidylserine in a Ca-dependen fashion.
In combination with RBS B0032 there is no detectable expression of annexin V.
RBS.4 (weaker) -- derivative of BBa_0030
Weaker RBS based on Ron Weiss thesis. Strengths relative to <bb_part>BBa_B0030</bb_part>, <bb_part>BBa_B0031</bb_part>, <bb_part>BBa_B0032</bb_part>.
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix ("RBS-3" in figure 4-14 of thesis). 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: Brian Chow
DNA is double-stranded. To translate genes of both strands, you need RBSs in both strands.
Please refer to the original (BB_B0034).
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene V message (BBa_M13005). The part aligns with base pairs 827-842 in M13K07 genome. It was identified as the RBS for the gene V message by Wezenbeek et al in Gene (1980) 11: 129-148
This RBS is part of the bacteriophage M13 genome, initiating translation of the gene VI message (BBa_M13006). The part aligns with base pairs 2840-2855 in M13K07 genome. It was identified as the RBS for gene VI message by Wezenbeek et al in Gene (1980) 11: 129-148