LOCUS       62056_11765        7268 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pGLM102(P_EF1a-tTA--Cer), complete sequence.
ACCESSION   MT891337
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7268)
  AUTHORS   Frei T, Cella F, Tedeschi F, Gutierrez J, Stan GB, Khammash M, 
            Siciliano V.
  TITLE     Characterization and mitigation of gene expression burden in 
            mammalian cells
  JOURNAL   Nat Commun 11 (1), 4641 (2020)
  PUBMED    32934213
REFERENCE   2  (bases 1 to 7268)
  AUTHORS   Frei T, Cella F, Tedeschi F, Gutierrez J, Stan G-B., Khammash M, 
            Siciliano V.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-AUG-2020) Synthetic and Systems Biology for 
            Biomedicine, Italian Institute of Technology-IIT, Largo Barsanti e 
            Martteucci 53, Naples, Campania 80125, Italy
REFERENCE   3  (bases 1 to 7268)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun"; date: "2020"; volume: "11"; issue: "1"; pages: "4641"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (15-AUG-2020) Synthetic and Systems Biology for Biomedicine, Italian
            Institute of Technology-IIT, Largo Barsanti e Martteucci 53, Naples,
            Campania 80125, Italy"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7268
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      1..266
                     /note="misc_signal; INS"
                     /regulatory_class="other"
     polyA_signal    283..507
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        529..1710
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation
                     factor EF-1-alpha"
     CDS             1741..2484
                     /label=tTA-Advanced
                     /note="improved tetracycline-controlled transactivator"
     CDS             2491..3207
                     /label=mCerulean
                     /note="enhanced monomeric variant of CFP (Rizzo et al.,
                     2004)"
     misc_binding    3221..3242
                     /label=miR-FF4 binding site
                     /bound_moiety="miR-FF4"
                     /note="mir FF4 target"
     misc_binding    3243..3264
                     /label=miR-FF4 binding site
                     /bound_moiety="miR-FF4"
                     /note="mir FF4 target"
     misc_binding    3265..3286
                     /label=miR-FF4 binding site
                     /bound_moiety="miR-FF4"
                     /note="mir FF4 target"
     promoter        complement(3323..3341)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     polyA_signal    3499..3620
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     regulatory      3710..3999
                     /note="misc_signal; INS"
                     /regulatory_class="other"
     rep_origin      complement(4078..4666)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4840..5697)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(5698..5802)
                     /label=AmpR promoter
     promoter        5872..6188
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             6285..6881
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
     regulatory      7114..7195
                     /label=SV40 pA
                     /note="SV40 pA"
                     /regulatory_class="polyA_signal_sequence"
ORIGIN
        1 gtcttctcca tcggatcctg atcaggggag ctcacgggga cagccccccc ccaaagcccc
       61 cagggatgta attacgtccc tcccccgcta gggggcagca gcgaccgccc ggggctccgc
      121 tccggtccgg cgctcccccc gcatcccgag ccggcagcgt gcggggacag cccgggcacg
      181 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
      241 acacctgggg gatacgggga aaaccgcggc atatggcctc gactgtgcct tctagttgcc
      301 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
      361 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
      421 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
      481 atgctgggga tgcggtgggc tctatggctt ctgagggcta gcatcgatgc tccggtgccc
      541 gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga ggggtcggca
      601 attgaaccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact
      661 ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta gtcgccgtga
      721 acgttctttt tcgcaacggg tttgccgcca gaacacaggt aagtgccgtg tgtggttccc
      781 gcgggcctgg cctctttacg ggttatggcc cttgcgtgcc ttgaattact tccacgcccc
      841 tggctgcagt acgtgattct tgatcccgag cttcgggttg gaagtgggtg ggagagttcg
      901 aggccttgcg cttaaggagc cccttcgcct cgtgcttgag ttgaggcctg gcctgggcgc
      961 tggggccgcc gcgtgcgaat ctggtggcac cttcgcgcct gtctcgctgc tttcgataag
     1021 tctctagcca tttaaaattt ttgatgacct gctgcgacgc tttttttctg gcaagatagt
     1081 cttgtaaatg cgggccaaga tctgcacact ggtatttcgg tttttggggc cgcgggcggc
     1141 gacggggccc gtgcgtccca gcgcacatgt tcggcgaggc ggggcctgcg agcgcggcca
     1201 ccgagaatcg gacgggggta gtctcaagct ggccggcctg ctctggtgcc tggcctcgcg
     1261 ccgccgtgta tcgccccgcc ctgggcggca aggctggccc ggtcggcacc agttgcgtga
     1321 gcggaaagat ggccgcttcc cggccctgct gcagggagct caaaatggag gacgcggcgc
     1381 tcgggagagc gggcgggtga gtcacccaca caaaggaaaa gggcctttcc gtcctcagcc
     1441 gtcgcttcat gtgactccac ggagtaccgg gcgccgtcca ggcacctcga ttagttctcg
     1501 agcttttgga gtacgtcgtc tttaggttgg ggggaggggt tttatgcgat ggagtttccc
     1561 cacactgagt gggtggagac tgaagttagg ccagcttggc acttgatgta attctccttg
     1621 gaatttgccc tttttgagtt tggatcttgg ttcattctca agcctcagac agtggttcaa
     1681 agtttttttc ttccatttca ggtgtcgtga cataaactgc cctagattaa ttaagccacc
     1741 atgtctagac tggacaagag caaagtcata aactctgctc tggaattact caatgaagtc
     1801 ggtatcgaag gcctgacgac aaggaaactc gctcaaaagc tgggagttga gcagcctacc
     1861 ctgtactggc acgtgaagaa caagcgggcc ctgctcgatg ccctggcaat cgagatgctg
     1921 gacaggcatc atacccactt ctgccccctg gaaggcgagt catggcaaga ctttctgcgg
     1981 aacaacgcca agtcattccg ctgtgctctc ctctcacatc gcgacggggc taaagtgcat
     2041 ctcggcaccc gcccaacaga gaaacagtac gaaaccctgg aaaatcagct cgcgttcctg
     2101 tgtcagcaag gcttctccct ggagaacgca ctgtacgctc tgtccgccgt gggccacttt
     2161 acactgggct gcgtattgga ggatcaggag catcaagtag caaaagagga aagagagaca
     2221 cctaccaccg attctatgcc cccacttctg agacaagcaa ttgagctgtt cgaccatcag
     2281 ggagccgaac ctgccttcct tttcggcctg gaactaatca tatgtggcct ggagaaacag
     2341 ctaaagtgcg aaagcggcgg gccggccgac gcccttgacg attttgactt agacatgctc
     2401 ccagccgatg cccttgacga ctttgacctt gatatgctgc ctgctgacgc tcttgacgat
     2461 tttgaccttg acatgctccc cgggggtacc atggtgagca agggcgagga gctgttcacc
     2521 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
     2581 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
     2641 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgacctg gggcgtgcag
     2701 tgcttcgccc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     2761 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     2821 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     2881 ttcaaggagg acggcaacat cctggggcac aagctggagt acaacgccat cagcgacaac
     2941 gtctatatca ccgccgacaa gcagaagaac ggcatcaagg ccaacttcaa gatccgccac
     3001 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     3061 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccaa gctgagcaaa
     3121 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     3181 actctcggca tggacgagct gtacaagtaa tgactccgga ccgcttgaag tctttaatta
     3241 aaccgcttga agtctttaat taaaccgctt gaagtcttta attaaaacta gtatgtacaa
     3301 gtagtaatct agagggccct attctatagt gtcacctaaa tgctagagct cgctgatcag
     3361 tccgtcgacg gatccaccgg atctagataa ctgatcataa tcagccatac cacatttgta
     3421 gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg
     3481 aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     3541 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3601 aaactcatca atgtatctta acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc
     3661 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gggagctcac
     3721 ggggacagcc cccccccaaa gcccccaggg atgtaattac gtccctcccc cgctaggggg
     3781 cagcagcgac cgcccggggc tccgctccgg tccggcgctc cccccgcatc ccgagccggc
     3841 agcgtgcggg gacagcccgg gcacggggaa ggtggcacgg gatcgctttc ctctgaacgc
     3901 ttctcgctgc tctttgagcc tgcagacacc tgggggatac ggggaaaatg atcaagatct
     3961 tctacactgg gacgcaagga ttatcctcga ggcgcgcgaa gaatcagggg ataacgcagt
     4021 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4081 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4141 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4201 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4261 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4321 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4381 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4441 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4501 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4561 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4621 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4681 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4741 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4801 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     4861 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     4921 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     4981 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5041 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5101 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5161 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5221 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     5281 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     5341 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5401 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caacacggga
     5461 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5521 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5581 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5641 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5701 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5761 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     5821 gccacctgga ggttttcacc gtcatcaccg aaacgcgcga gacgaaagat tggtgtggaa
     5881 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     5941 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     6001 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     6061 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     6121 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     6181 tttgcaaaaa gctagcttgc atgcctgcag gtcggccgcc acgaccggtg ccgccaccat
     6241 cccctgaccc acgcccctga cccctcacaa ggagacgacc ttccatgacc gagtacaagc
     6301 ccacggtgcg cctcgccacc cgcgacgacg tcccccgggc cgtacgcacc ctcgccgccg
     6361 cgttcgccga ctaccccgcc acgcgccaca ccgtcgaccc ggaccgccac atcgagcggg
     6421 tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg
     6481 tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg
     6541 cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc
     6601 agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg
     6661 ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc
     6721 ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc
     6781 gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg
     6841 aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgacgcccg ccccacgacc
     6901 cgcagcgccc gaccgaaagg agcgcacgac cccatggctc cgaccgaagc cacccggggc
     6961 ggccccgccg accccgcacc cgcccccgag gcccaccgac tctagaggat cataatcagc
     7021 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
     7081 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
     7141 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcaatct
     7201 cgtgatacgc ctatttttat aggttaatgt catggatgcc gcatagttaa gccagtatct
     7261 gctccctg
//
