LOCUS       62056_3865        4596 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pBI-CMV-EGFP-mKate-3xmiR31ts5'UTR, complete sequence.
ACCESSION   MT891345
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4596)
  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 4596)
  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 4596)
  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..4596
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    complement(22..103)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(223..939)
                     /label=EGFP
                     /note="enhanced GFP"
     regulatory      complement(963..1031)
                     /label=minimal CMV promoter 2
                     /note="minimal CMV promoter 2"
                     /regulatory_class="promoter"
     enhancer        1098..1401
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1402..1605
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     misc_feature    1659..1721
                     /label=3xmiR31-TS
                     /note="3xmiR31-TS"
     CDS             1735..2445
                     /label=PAmKate
                     /note="monomeric photoactivatable red fluorescent protein 
                     (Gunewardene et al., 2011)"
     CDS             2446..2463
                     /label=6xHis
                     /note="6xHis affinity tag"
     polyA_signal    2598..2679
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(2861..3449)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3623..4480)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(4481..4585)
                     /label=AmpR promoter
ORIGIN
        1 gaaaagtgcc acctgacgtc ggcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc
       61 tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat
      121 tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct
      181 ctacaaatgt ggtatggctg attatgatcc tctagactgc agcttgtaca gctcgtccat
      241 gccgagagtg atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc
      301 gttggggtct ttgctcaggg cggactgggt gctcaggtag tggttgtcgg gcagcagcac
      361 ggggccgtcg ccgatggggg tgttctgctg gtagtggtcg gcgagctgca cgctgccgtc
      421 ctcgatgttg tggcggatct tgaagttcac cttgatgccg ttcttctgct tgtcggccat
      481 gatatagacg ttgtggctgt tgtagttgta ctccagcttg tgccccagga tgttgccgtc
      541 ctccttgaag tcgatgccct tcagctcgat gcggttcacc agggtgtcgc cctcgaactt
      601 cacctcggcg cgggtcttgt agttgccgtc gtccttgaag aagatggtgc gctcctggac
      661 gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt cggggtagcg
      721 gctgaagcac tgcacgccgt aggtcagggt ggtcacgagg gtgggccagg gcacgggcag
      781 cttgccggtg gtgcagatga acttcagggt cagcttgccg taggtggcat cgccctcgcc
      841 ctcgccggac acgctgaact tgtggccgtt tacgtcgccg tccagctcga ccaggatggg
      901 caccaccccg gtgaacagct cctcgccctt gctcaccatg gtggcaccgg tgaattctcc
      961 aggcgatctg acggttcact aaacgagctc tgcttatata ggcctcccac cgtacacgcc
     1021 acctcgacat actcgagtag ttattaatag taatcaatta cggggtcatt agttcatagc
     1081 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     1141 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     1201 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     1261 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     1321 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
     1381 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
     1441 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     1501 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
     1561 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt
     1621 cagatccgct agggatccct tccatttcag gtggatccag ctatgccagc atcttgccta
     1681 gctatgccag catcttgcct agctatgcca gcatcttgcc tgctagccgc caccatggtg
     1741 tctaagggcg aagagctgat taaggagaac atgcacatga agctgtacat ggagggcacc
     1801 gtgaacaacc accacttcaa gtgcacatcc gagggcgaag gcaagcccta cgagggcacc
     1861 cagaccatga gaatcaaggt ggtcgagggc ggccctctcc ccttcgcctt cgacatcctg
     1921 gctaccagct tcatgtacgg cagcaaaacc ttcatcaacc acacccaggg catccccgac
     1981 ttctttaagc agtccttccc tgagggcttc acatgggaga gagtcaccac atacgaagac
     2041 gggggcgtgc tgaccgctac ccaggacacc agcctccagg acggctgcct catctacaac
     2101 gtcaagatca gaggggtgaa cttcccatcc aacggccctg tgatgcagaa gaaaacactc
     2161 ggctgggagg cctccaccga gatgctgtac cccgctgacg gcggcctgga aggcagaagc
     2221 gacatggccc tgaagctcgt gggcgggggc cacctgatct gcaacttgaa gaccacatac
     2281 agatccaaga aacccgctaa gaacctcaag atgcccggcg tctactatgt ggacagaaga
     2341 ctggaaagaa tcaaggaggc cgacaaagag acctacgtcg agcagcacga ggtggctgtg
     2401 gccagatact gcgacctccc tagcaaactg gggcacaaac ttaatcacca tcaccatcac
     2461 cattgaaagc ttgtcgacga tatctccaga ggatcataat cagccatacc acatttgtag
     2521 aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga
     2581 atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     2641 gcatcacaaa tttcacaaat aaagcatttt tttcactgcc ccgagcttcc tcgctcactg
     2701 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     2761 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     2821 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     2881 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     2941 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     3001 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     3061 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     3121 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     3181 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     3241 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3301 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3361 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3421 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3481 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     3541 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     3601 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     3661 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     3721 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     3781 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     3841 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     3901 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     3961 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     4021 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     4081 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     4141 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     4201 gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg ggataatacc gcgccacata
     4261 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     4321 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     4381 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     4441 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     4501 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4561 aaaataaaca aataggggtt ccgcgcacat ttcccc
//
