LOCUS       62056_20400        4137 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pT-GTW6-CMV-mKate_2xmiR31ts5'UTR, complete sequence.
ACCESSION   MT891349
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4137)
  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 4137)
  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 4137)
  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..4137
                     /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..779
                     /label=2xmiR31-TS
                     /note="2xmiR31-TS"
     CDS             793..1503
                     /label=PAmKate
                     /note="monomeric photoactivatable red fluorescent protein 
                     (Gunewardene et al., 2011)"
     CDS             1504..1521
                     /label=6xHis
                     /note="6xHis affinity tag"
     protein_bind    complement(1541..1565)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    1694..1815
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1822..2277)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2304..2408
                     /label=AmpR promoter
     CDS             2409..3266
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      3440..4028
                     /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 atcttgcctg
      781 ctagccgcca ccatggtgtc taagggcgaa gagctgatta aggagaacat gcacatgaag
      841 ctgtacatgg agggcaccgt gaacaaccac cacttcaagt gcacatccga gggcgaaggc
      901 aagccctacg agggcaccca gaccatgaga atcaaggtgg tcgagggcgg ccctctcccc
      961 ttcgccttcg acatcctggc taccagcttc atgtacggca gcaaaacctt catcaaccac
     1021 acccagggca tccccgactt ctttaagcag tccttccctg agggcttcac atgggagaga
     1081 gtcaccacat acgaagacgg gggcgtgctg accgctaccc aggacaccag cctccaggac
     1141 ggctgcctca tctacaacgt caagatcaga ggggtgaact tcccatccaa cggccctgtg
     1201 atgcagaaga aaacactcgg ctgggaggcc tccaccgaga tgctgtaccc cgctgacggc
     1261 ggcctggaag gcagaagcga catggccctg aagctcgtgg gcgggggcca cctgatctgc
     1321 aacttgaaga ccacatacag atccaagaaa cccgctaaga acctcaagat gcccggcgtc
     1381 tactatgtgg acagaagact ggaaagaatc aaggaggccg acaaagagac ctacgtcgag
     1441 cagcacgagg tggctgtggc cagatactgc gacctcccta gcaaactggg gcacaaactt
     1501 aatcaccatc accatcacca ttgattaatt aaagctagtt acccagcttt cttgtacaaa
     1561 gtggtacgcg tgacggccgc gactctagat cataatcagc cataccacat ttgtagaggt
     1621 tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc
     1681 aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     1741 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     1801 catcaatgta tcttaaggcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
     1861 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
     1921 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
     1981 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
     2041 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
     2101 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
     2161 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
     2221 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag
     2281 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     2341 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     2401 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     2461 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     2521 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     2581 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     2641 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     2701 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     2761 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     2821 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     2881 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     2941 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     3001 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     3061 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     3121 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     3181 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     3241 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     3301 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     3361 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     3421 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     3481 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     3541 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     3601 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     3661 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     3721 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     3781 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     3841 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     3901 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     3961 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     4021 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     4081 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcca
//
