BBa_J18951 1 BBa_J18951 FRB ~ mCitrine 2010-01-26T12:00:00Z 2015-08-31T04:08:36Z synthetic P19 expression cassette from Gruenberg et al. 2010 series of interaction reporter proteins. All proteins share the same architecture: * T7Start-domain1-5xGS-domain2-preSCsite-HisTag-T7Stop Only Domains1 and 2 vary and are taken from a list of protein interaction input and output (readout) parts. ===Expression=== Proteins were expressed in E. coli BL21 using T7 Polymerase and were purified by His-tag affinity. ===Reference=== * Gruenberg, Ferrar, van der Sloot, Serrano (2010): Building Blocks for Protein Interaction Devices false false _165_ 0 2175 165 It's complicated false Constructs were assembled according to R.F.C. 25. All parts were codon optimized for E. coli. false Raik Gruenberg component2066800 1 BBa_B0105 component2066794 1 BBa_J18919 component2066790 1 BBa_B0105 component2066782 1 BBa_J18912 component2066802 1 BBa_J18913 component2066788 1 BBa_J18922 component2066791 1 BBa_J18931 component2066785 1 BBa_J18926 component2066798 1 BBa_J18909 component2066793 1 BBa_B0105 component2066784 1 BBa_B0105 component2066787 1 BBa_B0105 component2066796 1 BBa_B0105 annotation2066796 1 BBa_B0105 range2066796 1 1155 1160 annotation2066798 1 BBa_J18909 range2066798 1 1161 1178 annotation2066800 1 BBa_B0105 range2066800 1 1179 1184 annotation2066793 1 BBa_B0105 range2066793 1 1125 1130 annotation2066784 1 BBa_B0105 range2066784 1 84 89 annotation2066785 1 BBa_J18926 range2066785 1 90 368 annotation2066791 1 BBa_J18931 range2066791 1 411 1124 annotation2066802 1 BBa_J18913 range2066802 1 1185 1319 annotation2066787 1 BBa_B0105 range2066787 1 369 374 annotation2066782 1 BBa_J18912 range2066782 1 1 83 annotation2066794 1 BBa_J18919 range2066794 1 1131 1154 annotation2066788 1 BBa_J18922 range2066788 1 375 404 annotation2066790 1 BBa_B0105 range2066790 1 405 410 BBa_J18912 1 T7start T7 promoter + RBS + START 2010-01-26T12:00:00Z 2015-08-31T04:08:36Z pET3a expression vector Strong T7 promoter and RBS including start codon for protein expression in E. coli or in-vitro. Part is provided in RFC 25 assembly format. false false _165_ 0 2175 165 It's complicated false * sequence is taken from the commonly used pET3a expression vector * deleted XbaI site between prom. and RBS * T7 promoter sequence (1-19) equals the one in BBa_I712074 * Conservative choice of part boundaries and modifications -- the part's size could probably be reduced. false Raik Gruenberg annotation2065586 1 start range2065586 1 81 83 BBa_J18926 1 FRB FRB(T2098L) 2010-01-26T12:00:00Z 2015-08-31T04:08:36Z human mTOR (aa sequence) gene synthesis Modified FKBP12-rapamycin-binding domain (FRB) of human mTOR Kinase (=mammalian Target of Rapamycin, also called FRAP = FKBP12-rapamycin associated protein). In presence of the immunosupressant drug rapamycin, FRB forms a high-affinity complex with FKBP12. There is no detectable FKBP12 - FRB interaction in absence of rapamycin. ===Modification for a non-toxic dimerizer=== Rapamycin arrests growth and proliferation of many cell types (reviewed in Jacinto and Hall, 2003) by inhibiting protein translation. The mutation T2089L (mTor numbering) makes FRB binding to a non-toxic Rapamycin analogue (rapalogue) AP21967 as well as the original Rapamycin. The FRB - FKBP12 heterodimerization can then be triggered by both Rapamycin and the non-toxic rapalogue. This variant of FRB and the synthetic dimerizer AP21967 are the basis of the Argent(tm) heterodimerization kit from Ariad Pharmaceuticals Inc. '''Note:''' Two additional mutations would convert FRB into FRB* which is used for conditional (drug-reversible) degradation of fusion proteins. These two additional mutations are not implemented in this part. ===Protein Parameters:=== * Number of amino acids: 93 * Molecular weight: 11285.9 * Theoretical pI: 6.47 false false _165_ 0 2175 165 It's complicated false * aa translation from human mTOR, * mutation T2089L (verified against pC4-RHE from Ariad) * GeneArt codon optimization for E. coli * gene synthesis false Raik Gruenberg BBa_J18909 1 HisTag His tag (6xHis; E. coli - optimized) 2010-01-15T12:00:00Z 2015-08-31T04:08:35Z Synthetic DNA Hexahistidine affinity tag. Binds zinc, nickel, or cobalt affinity resins. =Notes= This part is identical to BBa_I757013 (which does not seem to be available though). =References= Wikipedia article: [http://en.wikipedia.org/wiki/Polyhistidine-tag] false false _165_ 0 2175 165 It's complicated false Simple repetition of the His codon most frequent in E. coli -- a more balanced codon-choice may be better. false Raik Gruenberg annotation2065463 1 PolyHis range2065463 1 1 18 BBa_J18913 1 T7stop T7 terminator (incl. STOP) 2010-01-26T12:00:00Z 2015-08-31T04:08:36Z pET3a expression plasmid T7 terminator for protein expression in E. coli or in vitro. Part is provided in R.F.C. 25 assembly format. false false _165_ 0 2175 165 It's complicated false * taken from pET3a expression plasmid * added two STOP to 5' * conservative choice of part boundary, could probably be shortened false Raik Gruenberg annotation2065587 1 stop range2065587 1 1 6 BBa_J18922 1 5xGS 10aa [GS]x linker 2010-01-26T12:00:00Z 2015-08-31T04:08:36Z gene synthesis Highly flexible 10 amino acid linker. Translates to [GS]_5. Codon-optimize for E. coli, yeast, mammalian. false true _165_ 0 2175 165 It's complicated false * gene synthesis * codon optimized for E. coli * experimental distance distributions are described in Neuweiler et al. 2006 papers. false Raik Gruenberg BBa_J18931 1 mCitrine mCitrine 2010-01-26T12:00:00Z 2015-08-31T04:08:36Z gene synthesis Yellow fluorescent protein mCitrine. ===See also=== * BBa_J18930 ===References=== * Griesbeck O, Baird GS, Campbell RE, Zacharias DA, Tsien RY. (2001) Reducing the environmental sensitivity of yellow fluorescent protein. Mechanism and applications. PMID: 11387331 false false _165_ 0 2175 165 It's complicated false * starts with consensus VSKGEEL * ends with consensus DELYK false Raik Gruenberg BBa_B0105 1 Scar 25 RFC 25 Scar Sequence 2009-10-14T11:00:00Z 2015-08-31T04:07:21Z BBF RFC 25 This is the scar produced by assembly using RFC 25. If you are assembling a composite part using RFC 25, you can insert this part and specify blunt assembly to get the desired sequence. false false _1_ 0 25 397 Not in stock false Simple DNA sequence false Randy Rettberg annotation2041621 1 Scar 25 range2041621 1 1 6 BBa_J18919 1 preSCsite preScission protease cleavage site (E. coli-optimized) 2010-01-26T12:00:00Z 2015-08-31T04:08:36Z gene synthesis cleavage site for PreScission??? protease. preScission protease cleavage site (PreScission??? protease is a genetically engineered form of human rhinovirus 3C protease with a GST fusion, allowing for facile cleavage and purification of GST-tagged proteins along with protease removal after recombinant protein cleavage. Enables low-temperature cleavage of fusion proteins containing the eight-residue recognition sequence.) cleavage site: * Leu-Glu-Val-Leu-Phe-Gln↓Gly-Pro References ============== Walker et al. (1994): Efficient and rapid affinity purification of proteins using recombinant fusion proteases. PMID: 7764949 false false _165_ 0 2175 165 It's complicated false * gene synthesis * codon optimized for E. coli false Raik Gruenberg BBa_J18922_sequence 1 ggtagcggcagcggtagcggtagcggcagc BBa_B0105_sequence 1 accggc BBa_J18951_sequence 1 taatacgactcactatagggagaccacaacggtttccctctagcaataattttgtttaactttaagaaggagatatacatatgaccggcatcctgtggcatgaaatgtggcatgaaggcctggaagaagcgagccgtctgtattttggcgaacgtaacgtgaaaggcatgtttgaagtgctggaaccgctgcatgccatgatggaacgtggcccgcagaccctgaaagaaaccagctttaaccaggcgtatggccgtgatctgatggaagcgcaggaatggtgccgtaaatacatgaaaagcggcaacgtgaaagatctgctgcaagcgtgggatctgtattatcatgtgtttcgccgcatcagcaaaaccggcggtagcggcagcggtagcggtagcggcagcaccggcgtgagcaaaggcgaagaactgtttaccggcgtggtgccgattctggtggaactggatggtgatgtgaacggccataaatttagcgtgagcggcgaaggcgaaggtgatgcgacctatggcaaactgaccctgaaatttatttgcaccacgggtaaactgccggttccgtggccgaccctggtgaccacctttggctatggcctgatgtgctttgcgcgttatccggatcatatgaaacagcacgatttctttaaaagcgccatgccggaaggctatgtgcaggaacgcaccatcttttttaaagatgatggcaactataaaacccgtgcggaagtgaaatttgaaggcgataccctggtgaaccgtattgaactgaaaggcatcgatttcaaagaagatggcaacattctgggccataaactggaatataactacaacagccataacgtgtatatcatggcggataaacagaaaaacggcatcaaagtgaactttaaaatccgccacaacattgaagatggcagcgtgcagctggccgatcattatcagcagaacaccccgattggtgatggcccggtgctgctgccggataaccattatctgagctaccagagcaaactgagcaaagatccgaacgaaaaacgtgatcacatggtgctgctggaatttgtgaccgcagccggtattaccctgggcatggatgaactgtacaaaaccggcctggaagtgctgtttcagggcccgaccggccatcatcatcatcatcataccggctagtgactgctaacaaagcccgaaaggaagctgagttggctgctgccaccgctgagcaataactagcataaccccttggggcctctaaacgggtcttgaggggttttttgctgaaaggaggaactatatccggat BBa_J18913_sequence 1 tagtgactgctaacaaagcccgaaaggaagctgagttggctgctgccaccgctgagcaataactagcataaccccttggggcctctaaacgggtcttgaggggttttttgctgaaaggaggaactatatccggat BBa_J18909_sequence 1 catcatcatcatcatcat BBa_J18912_sequence 1 taatacgactcactatagggagaccacaacggtttccctctagcaataattttgtttaactttaagaaggagatatacatatg BBa_J18926_sequence 1 atcctgtggcatgaaatgtggcatgaaggcctggaagaagcgagccgtctgtattttggcgaacgtaacgtgaaaggcatgtttgaagtgctggaaccgctgcatgccatgatggaacgtggcccgcagaccctgaaagaaaccagctttaaccaggcgtatggccgtgatctgatggaagcgcaggaatggtgccgtaaatacatgaaaagcggcaacgtgaaagatctgctgcaagcgtgggatctgtattatcatgtgtttcgccgcatcagcaaa BBa_J18931_sequence 1 gtgagcaaaggcgaagaactgtttaccggcgtggtgccgattctggtggaactggatggtgatgtgaacggccataaatttagcgtgagcggcgaaggcgaaggtgatgcgacctatggcaaactgaccctgaaatttatttgcaccacgggtaaactgccggttccgtggccgaccctggtgaccacctttggctatggcctgatgtgctttgcgcgttatccggatcatatgaaacagcacgatttctttaaaagcgccatgccggaaggctatgtgcaggaacgcaccatcttttttaaagatgatggcaactataaaacccgtgcggaagtgaaatttgaaggcgataccctggtgaaccgtattgaactgaaaggcatcgatttcaaagaagatggcaacattctgggccataaactggaatataactacaacagccataacgtgtatatcatggcggataaacagaaaaacggcatcaaagtgaactttaaaatccgccacaacattgaagatggcagcgtgcagctggccgatcattatcagcagaacaccccgattggtgatggcccggtgctgctgccggataaccattatctgagctaccagagcaaactgagcaaagatccgaacgaaaaacgtgatcacatggtgctgctggaatttgtgaccgcagccggtattaccctgggcatggatgaactgtacaaa BBa_J18919_sequence 1 ctggaagtgctgtttcagggcccg 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