LOCUS       40924_43258        4976 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pTHSSe_37, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4976)
  AUTHORS   Segall-Shapiro TH, Sontag ED, Voigt CA.
  TITLE     Engineered promoters enable constant gene expression at any copy 
            number in bacteria
  JOURNAL   Nat. Biotechnol. (2018) In press
  PUBMED    29553576
REFERENCE   2  (bases 1 to 4976)
  AUTHORS   Segall-Shapiro TH, Sontag ED, Voigt CA.
  TITLE     Direct Submission
  JOURNAL   Submitted (06-MAR-2018) Synthetic Biology Center, Department of 
            Biological Engineering, Massachusetts Institute of Technology, 77 
            Massachusetts Avenue, Cambridge, MA 02139, USA
REFERENCE   3  (bases 1 to 4976)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 4976)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Biotechnol. (2018) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (06-MAR-2018) Synthetic Biology Center, Department of Biological 
            Engineering, Massachusetts Institute of Technology, 77 Massachusetts
            Avenue, Cambridge, MA 02139, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..4976
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..31
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     protein_bind    complement(37..56)
                     /label=lac operator (symmetric)
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     misc_RNA        73..123
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     regulatory      147..165
                     /note="B0032; derived from BBa_B0032"
                     /regulatory_class="ribosome_binding_site"
     CDS             166..879
                     /label=superfolder GFP
                     /note="GFP variant that folds robustly even when fused to
                     poorly folded proteins (Nager et al., 2011)"
     terminator      899..970
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      986..1013
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    1020..1040
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      1041..1098
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E.
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     CDS             complement(1259..2047)
                     /label=SmR
                     /note="aminoglycoside adenylyltransferase (Murphy, 1985)"
     rep_origin      complement(2642..3186)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin."
     terminator      complement(3676..3719)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     CDS             complement(3765..4844)
                     /label=lacI
                     /note="lac repressor"
     regulatory      complement(4845..4926)
                     /label=LacI promoter
                     /note="LacI promoter"
                     /regulatory_class="promoter"
ORIGIN
        1 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcgctc acaattctat
       61 ggactatgtt tagctgtcac cggatgtgct ttccggtctg atgagtccgt gaggacgaaa
      121 cagcctctac aaataatttt gtttaatcac acaggaaagt actagatgag caaaggagaa
      181 gaacttttca ctggagttgt cccaattctt gttgaattag atggtgatgt taatgggcac
      241 aaattttctg tccgtggaga gggtgaaggt gatgctacaa acggaaaact cacccttaaa
      301 tttatttgca ctactggaaa actacctgtt ccgtggccaa cacttgtcac tactctgacc
      361 tatggtgttc aatgcttttc ccgttatccg gatcacatga aacggcatga ctttttcaag
      421 agtgccatgc ccgaaggtta tgtacaggaa cgcactatat ctttcaaaga tgacgggacc
      481 tacaagacgc gtgctgaagt caagtttgaa ggtgataccc ttgttaatcg tatcgagtta
      541 aagggtattg attttaaaga agatggaaac attcttggac acaaactcga gtacaacttt
      601 aactcacaca atgtatacat cacggcagac aaacaaaaga atggaatcaa agctaacttc
      661 aaaattcgcc acaacgttga agatggttcc gttcaactag cagaccatta tcaacaaaat
      721 actccaattg gcgatggccc tgtcctttta ccagacaacc attacctgtc gacacaatct
      781 gtcctttcga aagatcccaa cgaaaagcgt gaccacatgg tccttcttga gtttgtaact
      841 gctgctggga ttacacatgg catggatgag ctctacaaat aatactagag ccaggcatca
      901 aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt
      961 gaacgctctc tactagagtc acactggctc accttcgggt gggcctttct gcgtttatat
     1021 actagtagcg gccgctgcag tccggcaaaa aagggcaagg tgtcaccacc ctgccctttt
     1081 tctttaaaac cgaaaagatt acttcgcgtt atgcaggctt cctcgctcac tgactcgctg
     1141 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatctcgagt
     1201 cccgtcaagt cagcgtaatg ctctgccagt gttacaacca attaaccaat tctgattatt
     1261 tgccgactac cttggtgatc tcgcctttca cgtagtggac aaattcttcc aactgatctg
     1321 tgcgcgaggc caagcgatct tcttcttgtc caagataagc ctgtctagct tcaagtatga
     1381 cgggctgata ctgggccggc aggcgctcca ttgcccagtc ggcagcgaca tccttcggcg
     1441 cgattttgcc ggttactgcg ctgtaccaaa tgcgggacaa cgtaagcact acatttcgct
     1501 catcgccagc ccagtcgggc ggcgagttcc atagcgttaa ggtttcattt agcgcctcaa
     1561 atagatcctg ttcaggaacc ggatcaaaga gttcctccgc cgctggacct accaaggcaa
     1621 cgctatgttc tcttgctttt gtcagcaaga tagccagatc aatgtcgatc gtggctggct
     1681 cgaagatacc tgcaagaatg tcattgcgct gccattctcc aaattgcagt tcgcgcttag
     1741 ctggataacg ccacggaatg atgtcgtcgt gcacaacaat ggtgacttct acagcgcgga
     1801 gaatctcgct ctctacaggg gaagccgaag tttccaaaag gtcgttgatc aaagctcgcc
     1861 gcgttgtttc atcaagcctt acggtcaccg taaccagcaa atcaatatca ctgtgtggct
     1921 tcaggccgcc atccactgcg gagccgtaca aatgtacggc cagcaacgtc ggttcgagat
     1981 ggcgctcgat gacgccaact acctctgata gttgagtcga tacttcggcg atcaccgctt
     2041 ccctcataac accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgatg
     2101 atatattttt atcttgtgca atgtaacatc agagattttg agacacaacg tggctttgtt
     2161 gaataaatcg aacttttgct gagttgaagg atcagatcac gcatcttccc gacaacgcag
     2221 accgttccgt ggcaaagcaa aagttcaaaa tcaccaactg gtccacctac aacaaagctc
     2281 tcatcaaccg tggctccctc actttctggc tggatgatgg ggcgattcag gcctggtatg
     2341 agtcagcaac accttcttca cgaggcagac ctcagcgcta gcggagtgta tactggctta
     2401 ctatgttggc actgatgagg gtgtcagtga agtgcttcat gtggcaggag aaaaaaggct
     2461 gcaccggtgc gtcagcagaa tatgtgatac aggatatatt ccgcttcctc gctcactgac
     2521 tcgctacgct cggtcgttcg actgcggcga gcggaaatgg cttacgaacg gggcggagat
     2581 ttcctggaag atgccaggaa gatacttaac agggaagtga gagggccgcg gcaaagccgt
     2641 ttttccatag gctccgcccc cctgacaagc atcacgaaat ctgacgctca aatcagtggt
     2701 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc cctggcggct ccctcgtgcg
     2761 ctctcctgtt cctgcctttc ggtttaccgg tgtcattccg ctgttatggc cgcgtttgtc
     2821 tcattccacg cctgacactc agttccgggt aggcagttcg ctccaagctg gactgtatgc
     2881 acgaaccccc cgttcagtcc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     2941 acccggaaag acatgcaaaa gcaccactgg cagcagccac tggtaattga tttagaggag
     3001 ttagtcttga agtcatgcgc cggttaaggc taaactgaaa ggacaagttt tggtgactgc
     3061 gctcctccaa gccagttacc tcggttcaaa gagttggtag ctcagagaac cttcgaaaaa
     3121 ccgccctgca aggcggtttt ttcgttttca gagcaagaga ttacgcgcag accaaaacga
     3181 tctcaagaag atcatcttat taaggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     3241 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     3301 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     3361 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     3421 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     3481 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     3541 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattcca
     3601 tggtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat aaaaataggc
     3661 gtatcacgag gcagaatttc agataaaaaa aatccttagc tttcgctaag gatgatttct
     3721 ggtcggccaa ttcgcgctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt
     3781 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     3841 tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agactggcaa cagctgattg
     3901 cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc
     3961 aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct atcttcggta
     4021 tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg
     4081 cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc
     4141 tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt
     4201 tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga
     4261 cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg
     4321 accagatgct ccacgcccag tcgcgtaccg tcctcatggg agaaaataat actgttgatg
     4381 ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca
     4441 gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg
     4501 agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc
     4561 accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc
     4621 gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt
     4681 tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc
     4741 cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag
     4801 acaccggcat actctgcgac atcgtataac gttactggtt tcatattcac caccctgaat
     4861 tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggcg
     4921 cgccgcttcg tcaggccaca tagctttctt gttctgatcg gaacgatcgt tggctg
//
