LOCUS       62056_20430        4158 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pT-GTW6-CMV-mKate_3xmiR31ts5'UTR, complete sequence.
ACCESSION   MT891350
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4158)
  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 4158)
  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 4158)
  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..4158
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    59..79
                     /label=attB4
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     enhancer        151..454
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        455..658
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     protein_bind    689..713
                     /label=attB1
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    738..800
                     /label=3xmiR31-TS
                     /note="3xmiR31-TS"
     CDS             814..1524
                     /label=PAmKate
                     /note="monomeric photoactivatable red fluorescent protein 
                     (Gunewardene et al., 2011)"
     CDS             1525..1542
                     /label=6xHis
                     /note="6xHis affinity tag"
     protein_bind    complement(1562..1586)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    1715..1836
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1843..2298)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2325..2429
                     /label=AmpR promoter
     CDS             2430..3287
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      3461..4049
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 catttcaggt gatccagtta tttaatagtc aattacggca tatatctagc agtgatatca
       61 actttgtata gaaaagttgg ctccgaattc tagttattaa tagtaatcaa ttacggggtc
      121 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
      181 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
      241 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
      301 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
      361 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
      421 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
      481 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
      541 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
      601 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg
      661 tttagtgaac cgtcagatcg aattcgaccc aagtttgtac aaaaaagcag gctgattctt
      721 ccatttcagg tggatccagc tatgccagca tcttgcctag ctatgccagc atcttgccta
      781 gctatgccag catcttgcct gctagccgcc accatggtgt ctaagggcga agagctgatt
      841 aaggagaaca tgcacatgaa gctgtacatg gagggcaccg tgaacaacca ccacttcaag
      901 tgcacatccg agggcgaagg caagccctac gagggcaccc agaccatgag aatcaaggtg
      961 gtcgagggcg gccctctccc cttcgccttc gacatcctgg ctaccagctt catgtacggc
     1021 agcaaaacct tcatcaacca cacccagggc atccccgact tctttaagca gtccttccct
     1081 gagggcttca catgggagag agtcaccaca tacgaagacg ggggcgtgct gaccgctacc
     1141 caggacacca gcctccagga cggctgcctc atctacaacg tcaagatcag aggggtgaac
     1201 ttcccatcca acggccctgt gatgcagaag aaaacactcg gctgggaggc ctccaccgag
     1261 atgctgtacc ccgctgacgg cggcctggaa ggcagaagcg acatggccct gaagctcgtg
     1321 ggcgggggcc acctgatctg caacttgaag accacataca gatccaagaa acccgctaag
     1381 aacctcaaga tgcccggcgt ctactatgtg gacagaagac tggaaagaat caaggaggcc
     1441 gacaaagaga cctacgtcga gcagcacgag gtggctgtgg ccagatactg cgacctccct
     1501 agcaaactgg ggcacaaact taatcaccat caccatcacc attgattaat taaagctagt
     1561 tacccagctt tcttgtacaa agtggtacgc gtgacggccg cgactctaga tcataatcag
     1621 ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa
     1681 cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg
     1741 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     1801 tagttgtggt ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata
     1861 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1921 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1981 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     2041 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     2101 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     2161 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     2221 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     2281 cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     2341 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     2401 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     2461 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     2521 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     2581 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     2641 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     2701 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     2761 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     2821 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     2881 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     2941 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     3001 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     3061 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     3121 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     3181 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     3241 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     3301 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     3361 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     3421 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     3481 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     3541 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     3601 atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     3661 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     3721 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     3781 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     3841 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     3901 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     3961 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4021 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4081 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4141 ataaccgtat taccgcca
//
