LOCUS       40924_43263        6488 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pTHSSe_38, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6488)
  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 6488)
  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 6488)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6488)
  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..6488
                     /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="RBS32A; derived from BBa_B0032"
                     /regulatory_class="ribosome_binding_site"
     CDS             166..2397
                     /codon_start=1
                     /product="TALE1"
                     /label=TALE1
                     /protein_id="AVR55108.1"
                     /translation="MVDLRTLGYSQQQQEKIKPKVRSTVAQHHEALVGHGFTHAHIVAL
                     SQHPAALGTVAVKYQDMIAALPEATHEAIVGVGKQWSGARALEALLTVAGELRGPPLQL
                     DTGQLLKIAKRGGVTAVEAVHAWRNALTGAPLNLTPEQVVAIASNIGGKQALETVQRLL
                     PVLCQAHGLTPEQVVAIASHDGGKQALETVQRLLPVLCQAHGLTPEQVVAIASNNGGKQ
                     ALETVQRLLPVLCQAHGLTPEQVVAIASNIGGKQALETVQRLLPVLCQAHGLTPEQVVA
                     IASHDGGKQALETVQRLLPVLCQAHGLTPEQVVAIASNGGGKQALETVQRLLPVLCQAH
                     GLTPEQVVAIASHDGGKQALETVQRLLPVLCQAHGLTPEQVVAIASNIGGKQALETVQR
                     LLPVLCQAHGLTPEQVVAIASHDGGKQALETVQRLLPVLCQAHGLTPEQVVAIASNGGG
                     KQALETVQRLLPVLCQAHGLTPEQVVAIASNIGGKQALETVQRLLPVLCQAHGLTPEQV
                     VAIASNGGGKQALETVQRLLPVLCQAHGLTPEQVVAIASNIGGRPALESIVAQLSRPDP
                     ALAALTNDHLVALACLGGRPALDAVKKGLPHAPALIKRTNRRIPERTSHRVADHAQVVR
                     VLGFFQCHSHPAQAFDDAMTQFGMSRHGLLQLFRRVGVTELEARSGTLPPASQRWDRIL
                     QASGMKRAKPSPTSTQTPDQASLHAFADSLERDLDAPSPMHEGDQTRAS"
     terminator      2411..2482
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      2498..2525
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    2532..2552
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      2553..2610
                     /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(2771..3559)
                     /label=SmR
                     /note="aminoglycoside adenylyltransferase (Murphy, 1985)"
     rep_origin      complement(4154..4698)
                     /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(5188..5231)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     CDS             complement(5277..6356)
                     /label=lacI
                     /note="lac repressor"
     regulatory      complement(6357..6438)
                     /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 actaaatggt agatttaaga
      181 actttaggat attcacagca gcaacaggaa aagatcaagc ccaaagttag gtcgacagtc
      241 gcgcagcatc acgaagcgct ggttggtcat gggtttacac atgcccacat cgtagcctta
      301 tcgcagcacc ctgcagccct tggcacggtc gccgtcaagt accaggacat gattgcggcg
      361 ttgccggaag ccacacatga ggcgatcgtc ggtgtgggga aacagtggag cggagcccga
      421 gcgcttgagg ccctgttgac ggtcgcggga gagctgagag ggcctcccct tcagctggac
      481 acgggccagt tgctgaagat cgcgaagcgg ggaggagtca cggcggtcga ggcggtgcac
      541 gcgtggcgca atgcgctcac gggagcaccc ctcaacctga ccccagagca ggtcgtggca
      601 atcgcctcca acattggcgg gaaacaggca ctcgagactg tccagcgcct gcttcccgtg
      661 ctgtgccaag cgcacggact caccccagag caggtcgtgg cgatcgcaag ccacgacgga
      721 ggaaagcaag ccttggaaac agtacagagg ctgttgcctg tgctgtgcca agcgcacggc
      781 ctcaccccag agcaggtcgt ggcaatcgcg agcaataacg gcggaaaaca ggctttggaa
      841 acggtgcaga ggctccttcc agtgctgtgc caagcgcacg gattaacccc agagcaggtc
      901 gtggcaatcg cctccaacat tggcgggaaa caggcactcg agactgtcca gcgcctgctt
      961 cccgtgctgt gccaagcgca cggcttaacc ccagagcagg tcgtggcgat cgcaagccac
     1021 gacggaggaa agcaagcctt ggaaacagta cagaggctgt tgcctgtgct gtgccaagcg
     1081 cacggactta ccccagagca ggtcgtggcc attgcctcga atggaggggg caaacaggcg
     1141 ttggaaaccg tacaacgatt gctgccggtg ctgtgccaag cgcacggcct taccccagag
     1201 caggtcgtgg cgatcgcaag ccacgacgga ggaaagcaag ccttggaaac agtacagagg
     1261 ctgttgcctg tgctgtgcca agcgcacgga ctaaccccag agcaggtcgt ggcaatcgcc
     1321 tccaacattg gcgggaaaca ggcactcgag actgtccagc gcctgcttcc cgtgctgtgc
     1381 caagcgcacg ggctcacccc agagcaggtc gtggcgatcg caagccacga cggaggaaag
     1441 caagccttgg aaacagtaca gaggctgttg cctgtgctgt gccaagcgca cgggctaacc
     1501 ccagagcagg tcgtggccat tgcctcgaat ggagggggca aacaggcgtt ggaaaccgta
     1561 caacgattgc tgccggtgct gtgccaagcg cacggcctaa ccccagagca ggtcgtggca
     1621 atcgcctcca acattggcgg gaaacaggca ctcgagactg tccagcgcct gcttcccgtg
     1681 ctgtgccaag cgcacgggtt aaccccagag caggtcgtgg ccattgcctc gaatggaggg
     1741 ggcaaacagg cgttggaaac cgtacaacga ttgctgccgg tgctgtgcca agcgcacgga
     1801 ctcacgcctg agcaggtagt ggctattgca tccaatatcg ggggcagacc cgcactggag
     1861 tcaatcgtgg cccagctttc gaggccggac cccgcgctgg ccgcactcac taatgatcat
     1921 cttgtagcgc tggcctgcct cggcggacga cccgccttgg atgcggtgaa gaaggggctc
     1981 ccgcacgcgc ctgcattgat taagcggacc aacagaagga ttcccgagag gacatcacat
     2041 cgagtggcag atcacgcgca agtggtccgc gtgctcggat tcttccagtg tcactcccac
     2101 cccgcacaag cgttcgatga cgccatgact caatttggta tgtcgagaca cggactgctg
     2161 cagctctttc gtagagtcgg tgtcacagaa ctcgaggccc gctcgggcac actgcctccc
     2221 gcctcccagc ggtgggacag gattctccaa gcgagcggta tgaaacgcgc gaagccttca
     2281 cctacgtcaa ctcagacacc tgaccaggcg agccttcatg cgttcgcaga ctcgctggag
     2341 agggatttgg acgcgccctc gcccatgcat gaaggggacc aaactcgcgc gtcataatag
     2401 gtccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     2461 ttgtttgtcg gtgaacgctc tctactagag tcacactggc tcaccttcgg gtgggccttt
     2521 ctgcgtttat atactagtag cggccgctgc agtccggcaa aaaagggcaa ggtgtcacca
     2581 ccctgccctt tttctttaaa accgaaaaga ttacttcgcg ttatgcaggc ttcctcgctc
     2641 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     2701 gtaatctcga gtcccgtcaa gtcagcgtaa tgctctgcca gtgttacaac caattaacca
     2761 attctgatta tttgccgact accttggtga tctcgccttt cacgtagtgg acaaattctt
     2821 ccaactgatc tgtgcgcgag gccaagcgat cttcttcttg tccaagataa gcctgtctag
     2881 cttcaagtat gacgggctga tactgggccg gcaggcgctc cattgcccag tcggcagcga
     2941 catccttcgg cgcgattttg ccggttactg cgctgtacca aatgcgggac aacgtaagca
     3001 ctacatttcg ctcatcgcca gcccagtcgg gcggcgagtt ccatagcgtt aaggtttcat
     3061 ttagcgcctc aaatagatcc tgttcaggaa ccggatcaaa gagttcctcc gccgctggac
     3121 ctaccaaggc aacgctatgt tctcttgctt ttgtcagcaa gatagccaga tcaatgtcga
     3181 tcgtggctgg ctcgaagata cctgcaagaa tgtcattgcg ctgccattct ccaaattgca
     3241 gttcgcgctt agctggataa cgccacggaa tgatgtcgtc gtgcacaaca atggtgactt
     3301 ctacagcgcg gagaatctcg ctctctacag gggaagccga agtttccaaa aggtcgttga
     3361 tcaaagctcg ccgcgttgtt tcatcaagcc ttacggtcac cgtaaccagc aaatcaatat
     3421 cactgtgtgg cttcaggccg ccatccactg cggagccgta caaatgtacg gccagcaacg
     3481 tcggttcgag atggcgctcg atgacgccaa ctacctctga tagttgagtc gatacttcgg
     3541 cgatcaccgc ttccctcata acaccccttg tattactgtt tatgtaagca gacagtttta
     3601 ttgttcatga tgatatattt ttatcttgtg caatgtaaca tcagagattt tgagacacaa
     3661 cgtggctttg ttgaataaat cgaacttttg ctgagttgaa ggatcagatc acgcatcttc
     3721 ccgacaacgc agaccgttcc gtggcaaagc aaaagttcaa aatcaccaac tggtccacct
     3781 acaacaaagc tctcatcaac cgtggctccc tcactttctg gctggatgat ggggcgattc
     3841 aggcctggta tgagtcagca acaccttctt cacgaggcag acctcagcgc tagcggagtg
     3901 tatactggct tactatgttg gcactgatga gggtgtcagt gaagtgcttc atgtggcagg
     3961 agaaaaaagg ctgcaccggt gcgtcagcag aatatgtgat acaggatata ttccgcttcc
     4021 tcgctcactg actcgctacg ctcggtcgtt cgactgcggc gagcggaaat ggcttacgaa
     4081 cggggcggag atttcctgga agatgccagg aagatactta acagggaagt gagagggccg
     4141 cggcaaagcc gtttttccat aggctccgcc cccctgacaa gcatcacgaa atctgacgct
     4201 caaatcagtg gtggcgaaac ccgacaggac tataaagata ccaggcgttt cccctggcgg
     4261 ctccctcgtg cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc cgctgttatg
     4321 gccgcgtttg tctcattcca cgcctgacac tcagttccgg gtaggcagtt cgctccaagc
     4381 tggactgtat gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc ggtaactatc
     4441 gtcttgagtc caacccggaa agacatgcaa aagcaccact ggcagcagcc actggtaatt
     4501 gatttagagg agttagtctt gaagtcatgc gccggttaag gctaaactga aaggacaagt
     4561 tttggtgact gcgctcctcc aagccagtta cctcggttca aagagttggt agctcagaga
     4621 accttcgaaa aaccgccctg caaggcggtt ttttcgtttt cagagcaaga gattacgcgc
     4681 agaccaaaac gatctcaaga agatcatctt attaaggggt ctgacgctca gtggaacgaa
     4741 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     4801 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     4861 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     4921 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     4981 cccagtgctg caatgatacc gcgataccca cgctcaccgg ctccagattt atcagcaata
     5041 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5101 cagtctattc catggtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     5161 ataaaaatag gcgtatcacg aggcagaatt tcagataaaa aaaatcctta gctttcgcta
     5221 aggatgattt ctggtcggcc aattcgcgct aacttacatt aattgcgttg cgctcactgc
     5281 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     5341 ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagactggc
     5401 aacagctgat tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg
     5461 gtttgcccca gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag
     5521 ctatcttcgg tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac
     5581 tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg
     5641 ggaacgatgc cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag
     5701 tcgccttccc gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca
     5761 gccagacgca gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg
     5821 tgacccaatg cgaccagatg ctccacgccc agtcgcgtac cgtcctcatg ggagaaaata
     5881 atactgttga tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag
     5941 gcagcttcca cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg
     6001 acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct
     6061 accatcgaca ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca
     6121 atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt
     6181 ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct
     6241 tccacttttt cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg
     6301 gtctgataag agacaccggc atactctgcg acatcgtata acgttactgg tttcatattc
     6361 accaccctga attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc
     6421 cattcgatgg cgcgccgctt cgtcaggcca catagctttc ttgttctgat cggaacgatc
     6481 gttggctg
//
