BBa_B0010 1 BBa_B0010 T1 from E. coli rrnB 2003-11-19T12:00:00Z 2015-08-31T04:07:20Z Transcriptional terminator consisting of a 64 bp stem-loop. false false _1_ 0 24 7 In stock false true Randy Rettberg annotation7018 1 BBa_B0010 range7018 1 1 80 annotation4184 1 stem_loop range4184 1 12 55 BBa_K343007 1 BBa_K343007 Photosensor generator 2010-10-15T11:00:00Z 2015-05-08T01:12:08Z Source Sensory Rhodopsin II and HtrII domains were taken from the genome of Natronomonas Pharaonis. The Tar methyl accepting chemotaxis protein was taken from Salmonella Enterica Serovar Typhimurium. The synthetic linker and fusing of the HAMP domains of HtrII and StTar was done by the team behind the referenced article (1). Molecular mechanism of the photosensor The fusion,chimera-protein coupled to the chemotaxis pathway. Figure taken from Trivedi et al.(2) The fusion,chimera-protein coupled to the chemotaxis pathway. Figure taken from Trivedi et al.(2) The photosensor acts directly on the tumbling frequency by effecting E. Coli's normal chemotaxis pathway. When exposed to bluelight, the sensory rhodopsin II will absorb the photons and undergo a change in ultrastructure, which is being transduced through HtrII on to the Tar domain. This effects decreases the autophosphorylation of the CheA protein, which in turn again decreases the amount of phosphorylated CheY, which just means that less of it will get phosphorylated. If there is less of the CheY-P, then there is a smaller chance of one of these molecules binding to the flagellar motor and making it turn clockwise, thereby inducing a lowered tumbling frequency in the system. The photosensor can also act in the opposite way, inducing a higher tumbling rate in the bacteria. Which effect the sensor will have depends on where NpHtrII and StTar are fused in the HAMP domain. If the fusion contains 20 more basepair of the HtrII domain and 20 less of the Tar domain, the photosensor would have the opposite effect and would be increasing the autophosphorylation of CheA. Usage and parameters The part requires retinal to work in E.Coli. This can be achieved through adding retinal to the liquid growth medium and/or the plates. Currently we are doing experiments on wether the part also functions with an internal retinal source, ie retinal synthesis in E. Coli. On top of adding retinal the cells will have to be grown in the dark for at least two hours after the addition of retinal to the growth medium if you want to see an effect. This is so that the photosensor is not exposed to light before the experiments and will result in maximum output if exposed to blue light. Results Bacteria containing this part will exhibit a lowered tumbling rate when exposed to blue light (wavelengths around 350nm - 450nm). This was analysed with the help of video microscopy and the open source software "CellTrack". The individual cells trajectory was tracked and their speed measured. The tracking results are as follows: From left to right, trajectory of: E.Coli with photosensor exposed to blue light, E.Coli with photosensor exposed to red light and E.Coli Mg1655 Wildtype exposed to blue light: (Blue dots show the location of the cell in the given frame, so the number of dots equals the number of frames from the sample.) The phototaxic bacteria move more in a straight line when exposed to bluelight, as can be seen when comparing the trajectories of the thee bacteria given earlier. These were taken from a batch of 10 cells tracked per sample. false false _465_ 0 6250 9 It's complicated true Design Notes NpSopII-NpHtrII-StTar fusion-chimera protein sequence from Jung K-H, Spudich EN, Trivedi VD and Spudich JL (1). The first 224 amino acid residues come from the NpSopII gene, encoding a bluelight photon receptor with 15 residues removed at the C-terminal end. The following 9 amino acids are a linker. the last part is HtrII fused with Tsr from E.Coli at the HAMP domain. The complex' first 125 amino acid residues come from HtrII and the remaining 279 from Tsr. The constitutive promoter R0040 was chosen for its medium strength, so that there would be a rather large amount of the fusion, chimera protein expressed, but not so much that it would be lethal to the cell. The easy repression and inhibition of repression through TetR and tetracycline makes for an easily controlled expression if that is needed. The doible terminator B0015 was chosen for it's reliability and good reviews on the partsregistry. false Maria Mollerup component2103033 1 BBa_B0015 component2103026 1 BBa_K343003 component2103019 1 BBa_J13002 annotation2103026 1 BBa_K343003 range2103026 1 81 2045 annotation2103033 1 BBa_B0015 range2103033 1 2054 2182 annotation2103019 1 BBa_J13002 range2103019 1 1 74 BBa_K343003 1 PS NpSopII-NpHtrII-StTar (M-fusion) 2010-07-01T11:00:00Z 2015-05-08T01:12:08Z / / false false _465_ 0 6083 9 It's complicated true / false Christian Kurtzhals annotation2072337 1 HtrII range2072337 1 700 1074 annotation2072341 1 Stop codon range2072341 1 1963 1965 annotation2072336 1 Linker range2072336 1 673 699 annotation2072334 1 Start codon range2072334 1 1 3 annotation2072340 1 Tar range2072340 1 1075 1965 annotation2072335 1 Sensory Rhodopsin II range2072335 1 1 672 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 annotation7020 1 BBa_B0012 range7020 1 1 41 annotation1687 1 stop range1687 1 34 34 annotation1690 1 polya range1690 1 28 41 annotation1686 1 T7 TE range1686 1 8 27 BBa_J13002 1 BBa_J13002 TetR repressed POPS/RIPS generator 2005-06-15T11:00:00Z 2015-08-31T04:08:29Z Released HQ 2013 -- No description -- false true _37_5_ 0 88 37 In stock false true Jeff Tabor component1535786 1 BBa_B0034 component1535778 1 BBa_R0040 annotation1535778 1 BBa_R0040 range1535778 1 1 54 annotation1535786 1 BBa_B0034 range1535786 1 63 74 BBa_B0015 1 BBa_B0015 double terminator (B0010-B0012) 2003-07-16T11:00:00Z 2015-08-31T04:07:20Z Released HQ 2013 Double terminator consisting of BBa_B0010 and BBa_B0012 false true _1_ 0 24 7 In stock false true Reshma Shetty component1916612 1 BBa_B0012 component1916610 1 BBa_B0010 annotation1916612 1 BBa_B0012 range1916612 1 89 129 annotation1916610 1 BBa_B0010 range1916610 1 1 80 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_R0040 1 p(tetR) TetR repressible promoter 2003-01-31T12:00:00Z 2015-05-08T01:14:14Z Lutz, R., Bujard, H., <em>Nucleic Acids Research</em> (1997) 25, 1203-1210. Released HQ 2013 Sequence for pTet inverting regulator driven by the TetR protein.</P> false true _1_ 0 24 7 In stock false <P> <P>BBa_R0040 TetR-Regulated Promoter is based on a cI promoter. It has been modified to include two TetR binding sites and the BioBrick standard assembly head and tail restriction sites.<P> true June Rhee, Connie Tao, Ty Thomson, Louis Waldman annotation1986787 1 -10 range1986787 1 43 48 annotation1986786 1 TetR 2 range1986786 1 26 44 annotation1986783 1 TetR 1 range1986783 1 1 19 annotation1986784 1 BBa_R0040 range1986784 1 1 54 annotation1986785 1 -35 range1986785 1 20 25 BBa_B0010_sequence 1 ccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctc BBa_J13002_sequence 1 tccctatcagtgatagagattgacatccctatcagtgatagagatactgagcactactagagaaagaggagaaa BBa_B0034_sequence 1 aaagaggagaaa BBa_K343003_sequence 1 atggtgggacttacgaccctcttttggctcggcgcaatcggcatgctcgtcggcacgctcgcgttcgcgtgggccggccgtgacgccggaagcggcgagcgacggtactacgtgacacttgtcggcatcagtggtatcgcagcagtcgcctacgccgttatggcgctgggtgtcggctgggttcccgtggccgaacggactgttttcgtcccccggtacatcgactggattctcacaaccccgctcatcgtctacttcctcgggctgcttgcggggcttgatagtcgggagttcggcatcgtcatcacgctcaacaccgtggtcatgctcgccggcttcgccggggcgatggtgcccggtatcgagcgctacgcgctgttcggcatgggggcggtcgcattcatcggactggtctactacctcgtcgggccgatgaccgaaagcgccagccagcggtcctccggaatcaagtcgctgtacgtccgcctccgaaacctgacggtcgtcctctgggcgatttatccgttcatctggctgcttggaccgccgggcgtggcgctgctgacaccgactgtcgacgtggcgcttatcgtctaccttgacctcgtcacgaaggtcgggttcggattcatcgcactcgatgctgcggcgacacttcgggccgaacacggagcgtcggcgtcgaacggcgcgtcggcgatgtcgctgaacgtatcacggctccttctccccggccgtgtccggcacagttatacggggaaaatgggtgccaccttcgcctttgtcggcgcgttaacggtgctcttcggagccatcgcatacggtgaggtaaccgccgccgccgcgaccggcgatgccgcagccgtacaggaggcggcagtatcggccattctcgggctcatcatcctgctcgggatcaacctcgggctcgttgctgccacgctgggcggtgacaccgccgcctcgctttcaacgctggccgcgaaggcctcgcggatgggcgacggcgacctcgatgtcgagcttgagacccgtcgcgaggacgaaatcggcgacctctatgcggccttcgaccaacgctcgctgattgacaccgtaacgcaggttcgtgaaggttcggatgcgatttattccggcaccagtgaaattgccgccggtaataccgacctctcttcccgtaccgaacagcaggcctccgctctggaggagacggctgccagcatggaacaactgacggccaccgtgaagcaaaacgccgataacgcccgccaggcttcgcaactggcgcaaagcgcctccgagaccgcgcgtcatggcggcaaagtggtcgacggcgtagtaaacactatgcacgaaattgccgacagttcgaaaaaaatcgctgacattatcagcgttatcgacggtattgccttccagactaacattctggcgctgaacgcggcggtagaagcggcgcgcgcgggagagcaggggcgcggttttgcggtcgtggcaggcgaggtgcgtaatctggccagccgcagcgcccaggcggcgaaagaaataaaagcgttgattgaagattccgtctcgcgtgtcgataccggttctgtgctggtggaaagcgccggggaaaccatgactgacatcgtcaatgccgttacgcgcgtcacggatatcatgggcgaaatcgcctccgcctcggatgagcaaagccggggtatcgatcaggtcgctttggccgtttccgaaatggatcgcgtaacgcaacagaacgcctcgctggttcaggaatccgcagcggccgccgccgcgctggaagagcaggccagccgtctgacccaggcggtatcggctttccgcctggcatcgcgaccgctggcggtaaataaacctgagatgcgtttgtcagtgaacgctcagtccggcaatacgccgcagtcattagccgccagggatgatgcgaactgggaaaccttctga BBa_R0040_sequence 1 tccctatcagtgatagagattgacatccctatcagtgatagagatactgagcac BBa_K343007_sequence 1 tccctatcagtgatagagattgacatccctatcagtgatagagatactgagcactactagagaaagaggagaaatactagatggtgggacttacgaccctcttttggctcggcgcaatcggcatgctcgtcggcacgctcgcgttcgcgtgggccggccgtgacgccggaagcggcgagcgacggtactacgtgacacttgtcggcatcagtggtatcgcagcagtcgcctacgccgttatggcgctgggtgtcggctgggttcccgtggccgaacggactgttttcgtcccccggtacatcgactggattctcacaaccccgctcatcgtctacttcctcgggctgcttgcggggcttgatagtcgggagttcggcatcgtcatcacgctcaacaccgtggtcatgctcgccggcttcgccggggcgatggtgcccggtatcgagcgctacgcgctgttcggcatgggggcggtcgcattcatcggactggtctactacctcgtcgggccgatgaccgaaagcgccagccagcggtcctccggaatcaagtcgctgtacgtccgcctccgaaacctgacggtcgtcctctgggcgatttatccgttcatctggctgcttggaccgccgggcgtggcgctgctgacaccgactgtcgacgtggcgcttatcgtctaccttgacctcgtcacgaaggtcgggttcggattcatcgcactcgatgctgcggcgacacttcgggccgaacacggagcgtcggcgtcgaacggcgcgtcggcgatgtcgctgaacgtatcacggctccttctccccggccgtgtccggcacagttatacggggaaaatgggtgccaccttcgcctttgtcggcgcgttaacggtgctcttcggagccatcgcatacggtgaggtaaccgccgccgccgcgaccggcgatgccgcagccgtacaggaggcggcagtatcggccattctcgggctcatcatcctgctcgggatcaacctcgggctcgttgctgccacgctgggcggtgacaccgccgcctcgctttcaacgctggccgcgaaggcctcgcggatgggcgacggcgacctcgatgtcgagcttgagacccgtcgcgaggacgaaatcggcgacctctatgcggccttcgaccaacgctcgctgattgacaccgtaacgcaggttcgtgaaggttcggatgcgatttattccggcaccagtgaaattgccgccggtaataccgacctctcttcccgtaccgaacagcaggcctccgctctggaggagacggctgccagcatggaacaactgacggccaccgtgaagcaaaacgccgataacgcccgccaggcttcgcaactggcgcaaagcgcctccgagaccgcgcgtcatggcggcaaagtggtcgacggcgtagtaaacactatgcacgaaattgccgacagttcgaaaaaaatcgctgacattatcagcgttatcgacggtattgccttccagactaacattctggcgctgaacgcggcggtagaagcggcgcgcgcgggagagcaggggcgcggttttgcggtcgtggcaggcgaggtgcgtaatctggccagccgcagcgcccaggcggcgaaagaaataaaagcgttgattgaagattccgtctcgcgtgtcgataccggttctgtgctggtggaaagcgccggggaaaccatgactgacatcgtcaatgccgttacgcgcgtcacggatatcatgggcgaaatcgcctccgcctcggatgagcaaagccggggtatcgatcaggtcgctttggccgtttccgaaatggatcgcgtaacgcaacagaacgcctcgctggttcaggaatccgcagcggccgccgccgcgctggaagagcaggccagccgtctgacccaggcggtatcggctttccgcctggcatcgcgaccgctggcggtaaataaacctgagatgcgtttgtcagtgaacgctcagtccggcaatacgccgcagtcattagccgccagggatgatgcgaactgggaaaccttctgatactagagccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata BBa_B0012_sequence 1 tcacactggctcaccttcgggtgggcctttctgcgtttata BBa_B0015_sequence 1 ccaggcatcaaataaaacgaaaggctcagtcgaaagactgggcctttcgttttatctgttgtttgtcggtgaacgctctctactagagtcacactggctcaccttcgggtgggcctttctgcgtttata 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