LOCUS       40924_43338        5979 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pTHSSe_69, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5979)
  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 5979)
  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 5979)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5979)
  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..5979
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      13..73
                     /label=L3S2P21
                     /note="L3S2P21"
                     /regulatory_class="terminator"
     regulatory      78..146
                     /label=PT7A1w2
                     /note="PT7A1w2"
                     /regulatory_class="promoter"
     ncRNA           147..225
                     /product="SarJ"
                     /label=ribozyme
                     /note="insulator"
                     /ncRNA_class="ribozyme"
     regulatory      226..245
                     /label=RBSsp1
                     /note="RBSsp1"
                     /regulatory_class="ribosome_binding_site"
     CDS             246..2885
                     /codon_start=1
                     /product="TALEsp1"
                     /label=TALEsp1
                     /protein_id="AVR55192.1"
                     /translation="MVDLRTLGYSQQQQEKIKPKVRSTVAQHHEALVGHGFTHAHIVAL
                     SQHPAALGTVAVKYQDMIAALPEATHEAIVGVGKQWSGARALEALLTVAGELRGPPLQL
                     DTGQLLKIAKRGGVTAVEAVHAWRNALTGAPLNLTPDQVVAIASNGGGKQALETVQRLL
                     PVLCQDHGLTPEQVVAIASNIGGKQALETVQRLLPVLCQAHGLTPDQVVAIASNGGGKQ
                     ALETVQRLLPVLCQAHGLTPAQVVAIASNIGGKQALETVQRLLPVLCQDHGLTPDQVVA
                     IASNIGGKQALETVQRLLPVLCQDHGLTPEQVVAIASHDGGKQALETVQRLLPVLCQAH
                     GLTPDQVVAIASNGGGKQALETVQRLLPVLCQAHGLTPAQVVAIASHDGGKQALETVQR
                     LLPVLCQDHGLTPDQVVAIASNIGGKQALETVQRLLPVLCQDHGLTPEQVVAIASNIGG
                     KQALETVQRLLPVLCQAHGLTPDQVVAIASHDGGKQALETVQRLLPVLCQAHGLTPAQV
                     VAIASHDGGKQALETVQRLLPVLCQDHGLTPDQVVAIASNGGGKQALETVQRLLPVLCQ
                     DHGLTPEQVVAIASNIGGKQALETVQRLLPVLCQAHGLTPDQVVAIASNGGGKQALETV
                     QRLLPVLCQAHGLTPAQVVAIASNIGGKQALETVQRLLPVLCQDHGLTPEQVVAIASNN
                     GGRPALESIVAQLSRPDPALAALTNDHLVALACLGGRPALDAVKKGLPHAPALIKRTNR
                     RIPERTSHRVADHAQVVRVLGFFQCHSHPAQAFDDAMTQFGMSRHGLLQLFRRVGVTEL
                     EARSGTLPPASQRWDRILQASGMKRAKPSPTSTQTPDQASLHAFADSLERDLDAPSPMH
                     EGDQTRAS"
     regulatory      complement(2891..2980)
                     /label=ECK120029600
                     /note="ECK120029600"
                     /regulatory_class="terminator"
     regulatory      2985..3059
                     /label=PUPsp1
                     /note="PUPsp1"
                     /regulatory_class="promoter"
     protein_bind    3017..3034
                     /label=TALEsp1 binding site
                     /bound_moiety="TALEsp1"
     misc_RNA        3061..3111
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     regulatory      3135..3156
                     /label=RBSPhlF
                     /note="RBSPhlF"
                     /regulatory_class="ribosome_binding_site"
     CDS             3157..3759
                     /codon_start=1
                     /product="PhlF"
                     /label=PhlF
                     /protein_id="AVR55193.1"
                     /translation="MARTPSRSSIGSLRSPHTHKAILTSTIEILKECGYSGLSIESVAR
                     RAGASKPTIYRWWTNKAALIAEVYENESEQVRKFPDLGSFKADLDFLLRNLWKVWRETI
                     CGEAFRCVIAEAQLDPATLTQLKDQFMERRREMPKKLVENAISNGELPKDTNRELLLDM
                     IFGFCWYRLLTEQLTVEQDIEEFTFLLINGVCPGTQR"
     terminator      3776..3847
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      3863..3890
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    3909..3929
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      3930..3987
                     /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."
     rep_origin      4159..4747
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(4954..5048)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(5072..5728)
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     promoter        complement(5729..5832)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     terminator      complement(5912..5955)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    5958..5979
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with
                     'ATG'"
ORIGIN
        1 gaagacacgg agctcggtac caaattccag aaaagaggcc tcccgaaagg ggggcctttt
       61 ttcgttttgg tcctactggc gcgcctcagt cagagtattg acttaaagtc taacctatag
      121 gaccgttaca gccatcgaga gctgcgagac tgtcgccgga tgtgtatccg acctgacgat
      181 ggcccaaaag ggccgaaaca gtcctctaca aataattttg tttaaatcaa ttcacctgcg
      241 tgaaaatggt agatttaaga actttaggat attcacagca gcaacaggaa aagatcaagc
      301 ccaaagttag gtcgacagtc gcgcagcatc acgaagcgct ggttggtcat gggtttacac
      361 atgcccacat cgtagcctta tcgcagcacc ctgcagccct tggcacggtc gccgtcaagt
      421 accaggacat gattgcggcg ttgccggaag ccacacatga ggcgatcgtc ggtgtgggga
      481 aacagtggag cggagcccga gcgcttgagg ccctgttgac ggtcgcggga gagctgagag
      541 ggcctcccct tcagctggac acgggccagt tgctgaagat cgcgaagcgg ggaggagtca
      601 cggcggtcga ggcggtgcac gcgtggcgca atgcgctcac gggagcaccc ctcaatctga
      661 ccccggatca ggtggttgcg atcgccagta atggcggagg caaacaagca ctcgaaactg
      721 tacagcgcct cctgccggta ctgtgccaag atcatggctt gacgcctgag caagtcgtag
      781 ctattgcatc aaacatcggt ggtaaacagg cgctggaaac cgtacaacga ttactccctg
      841 tcttatgcca ggcacacggt ctgacccccg accaagttgt agccattgcg tctaatggcg
      901 gcgggaaaca ggccctggaa acggtccaac gtctgttacc cgttctgtgt caggctcacg
      961 gtctgacccc tgcccaggta gttgcaattg ccagcaacat cggcgggaaa caagcgctgg
     1021 agactgtgca gcgtctgctc cctgtgttat gccaagatca tgggctcact ccggatcagg
     1081 tggtggccat cgcttccaat attggcggta aacaggcgct ggagacagtg caacgacttt
     1141 tacctgttct ctgccaggat catggtctaa ctcccgagca ggtcgtcgcc atcgcctctc
     1201 atgacggcgg gaaacaagcg ttggaaactg tccagcgact cctgccggtt ttgtgccagg
     1261 cccacgggct tactcctgac caggtagttg cgatcgcgtc aaatgggggt ggcaaacaag
     1321 ccctcgaaac cgtgcaacgc ctgctgcccg tcttgtgcca agctcatggg ctgactccgg
     1381 cgcaagtagt cgcgattgcg agccacgatg gcggtaagca ggcactggaa acggttcagc
     1441 gcctgctccc ggttctatgc caggatcacg gcctgacccc ggaccaggtc gtcgcgatcg
     1501 cgtcaaatat cggtggcaaa caagctttgg agacagtaca gcgcctgtta ccagtgcttt
     1561 gccaggacca tggtctgacc cctgagcaag tagtggcgat cgcttctaat attgggggca
     1621 aacaagcgct ggaaacagta cagcgtctgt taccggtcct atgccaggca catggcctga
     1681 cccctgatca ggtggtagcc attgccagtc atgatggcgg taaacaggcg cttgagactg
     1741 tccaacgtct gctgccggtc ctctgtcagg ctcatggcct gacgccagct caagtcgtgg
     1801 ctatcgcttc gcatgatggc ggaaaacagg cactggagac tgtgcagcga ctgttgccag
     1861 ttctgtgtca ggatcacggt ttaactccgg accaggtggt cgctattgcg tcgaatgggg
     1921 gcggtaaaca agcgctggaa actgtgcaac gtttgctccc agttctgtgc caggaccatg
     1981 ggctgactcc ggaacaggta gtggccattg cttctaatat tggtgggaaa caggcgctgg
     2041 aaaccgtgca gcgcctgctt ccagtgcttt gccaggccca tggcctgacg ccagatcagg
     2101 tggttgctat agccagcaat ggcggcggta aacaggccct cgaaaccgtc cagcgcctgc
     2161 tccctgtgct gtgccaggcc catgggctta ccccagcgca agtagtggcg attgcgtcta
     2221 atattggtgg taaacaggcg ttggagactg tacaacgcct gctgccagtt ttatgccaag
     2281 atcatggtct gacccctgag caggtagtgg ctattgcatc caacaacggg ggcagacccg
     2341 cactggagtc aatcgtggcc cagctttcga ggccggaccc cgcgctggcc gcactcacta
     2401 atgatcatct tgtagcgctg gcctgcctcg gcggacgacc cgccttggat gcggtgaaga
     2461 aggggctccc gcacgcgcct gcattgatta agcggaccaa cagaaggatt cccgagagga
     2521 catcacatcg agtggcagat cacgcgcaag tggtccgcgt gctcggattc ttccagtgtc
     2581 actcccaccc cgcacaagcg ttcgatgacg ccatgactca atttggtatg tcgagacacg
     2641 gactgctgca gctctttcgt agagtcggtg tcacagaact cgaggcccgc tcgggcacac
     2701 tgcctcccgc ctcccagcgg tgggacagga ttctccaagc gagcggtatg aaacgcgcga
     2761 agccttcacc tacgtcaact cagacacctg accaggcgag ccttcatgcg ttcgcagact
     2821 cgctggagag ggatttggac gcgccctcgc ccatgcatga aggggaccaa actcgcgcgt
     2881 cataataggt ttcagccaaa aaacttaaga ccgccggtct tgtccactac cttgcagtaa
     2941 tgcggtggac aggatcggcg gttttctttt ctcttctcaa gcttatcccg aaaatttatc
     3001 aaaaagagta ttgacttata actcaaccta taggatactt acagccatcg agagctgcga
     3061 gctgtcaccg gatgtgcttt ccggtctgat gagtccgtga ggacgaaaca gcctctacaa
     3121 ataattttgt ttaatactca ttggtgaaag tactagatgg cacgtacccc gagccgtagc
     3181 agcattggta gcctgcgtag tccgcatacc cataaagcaa ttctgaccag caccattgaa
     3241 atcctgaaag aatgtggtta tagcggtctg agcattgaaa gcgttgcacg tcgtgccggt
     3301 gcaagcaaac cgaccattta tcgttggtgg accaataaag cagcactgat tgccgaagtg
     3361 tatgaaaatg aaagcgaaca ggtgcgtaaa tttccggatc tgggtagctt taaagccgat
     3421 ctggattttc tgctgcgtaa tctgtggaaa gtttggcgtg aaaccatttg tggtgaagca
     3481 tttcgttgtg ttattgcaga agcacagctg gaccctgcaa ccctgaccca gctgaaagat
     3541 cagtttatgg aacgtcgtcg tgagatgccg aaaaaactgg ttgaaaatgc cattagcaat
     3601 ggtgaactgc cgaaagatac caatcgtgaa ctgctgctgg atatgatttt tggtttttgt
     3661 tggtatcgcc tgctgaccga acagctgacc gttgaacagg atattgaaga atttaccttc
     3721 ctgctgatta atggtgtttg tccgggtaca cagcgttaat actagagcca ggcatcaaat
     3781 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
     3841 cgctctctac tagagtcaca ctggctcacc ttcgggtggg cctttctgcg tttatacgct
     3901 atgtcttcta ctagtagcgg ccgctgcagt ccggcaaaaa agggcaaggt gtcaccaccc
     3961 tgcccttttt ctttaaaacc gaaaagatta cttcgcgtta tgcaggcttc ctcgctcact
     4021 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     4081 attcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     4141 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     4201 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     4261 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     4321 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     4381 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     4441 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     4501 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     4561 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     4621 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     4681 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     4741 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     4801 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtcgat tatcaaaaag
     4861 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     4921 tgagtaaact tggtctgaca gctcgaggct tggattctca ccaataaaaa acgcccggcg
     4981 gcaaccgagc gttctgaaca aatccagatg gagttctgag gtcattactg gatctatcaa
     5041 caggagtcca agcgagctcg atatcaaatt acgccccgcc ctgccactca tcgcagtact
     5101 gttgtaattc attaagcatt ctgccgacat ggaagccatc acaaacggca tgatgaacct
     5161 gaatcgccag cggcatcagc accttgtcgc cttgcgtata atatttgccc atggtgaaaa
     5221 cgggggcgaa gaagttgtcc atattggcca cgtttaaatc aaaactggtg aaactcaccc
     5281 agggattggc tgagacgaaa aacatattct caataaaccc tttagggaaa taggccaggt
     5341 tttcaccgta acacgccaca tcttgcgaat atatgtgtag aaactgccgg aaatcgtcgt
     5401 ggtattcact ccagagcgat gaaaacgttt cagtttgctc atggaaaacg gtgtaacaag
     5461 ggtgaacact atcccatatc accagctcac cgtctttcat tgccatacga aattccggat
     5521 gagcattcat caggcgggca agaatgtgaa taaaggccgg ataaaacttg tgcttatttt
     5581 tctttacggt ctttaaaaag gccgtaatat ccagctgaac ggtctggtta taggtacatt
     5641 gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat atatcaacgg
     5701 tggtatatcc agtgattttt ttctccattt tagcttcctt agctcctgaa aatctcgata
     5761 actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta
     5821 cgtgcccgat caactcgagt gccacctgac gtctaagaaa ccattattat catgacatta
     5881 acctataaaa ataggcgtat cacgaggcag aatttcagat aaaaaaaatc cttagctttc
     5941 gctaaggatg atttctggaa ttcgcggccg cttctagag
//
