LOCUS       62056_20375        4166 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pT-GTW6-CMV-mKate_2xmiR21ts3'UTR, complete sequence.
ACCESSION   MT891358
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4166)
  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 4166)
  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 4166)
  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..4166
                     /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    717..737
                     /label=attB1
                     /note="core recombination site for the Gateway(R) BP
                     reaction"
     CDS             775..792
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             793..1500
                     /label=PAmKate
                     /note="monomeric photoactivatable red fluorescent protein 
                     (Gunewardene et al., 2011)"
     misc_feature    1512..1555
                     /label=2xmiR21-TS
                     /note="2xmiR21-TS"
     protein_bind    complement(1570..1594)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    1723..1844
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1851..2306)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2333..2437
                     /label=AmpR promoter
     CDS             2438..3295
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      3469..4057
                     /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 cgtcagatca gagctggttt agtgaaccgt cagatcgaat tcgacccaag
      721 tttgtacaaa gaagcaggct gattcttcca tttcaggtgg atcccgccac catgcaccat
      781 caccatcacc atgtgtctaa gggcgaagag ctgattaagg agaacatgca catgaagctg
      841 tacatggagg gcaccgtgaa caaccaccac ttcaagtgca catccgaggg cgaaggcaag
      901 ccctacgagg gcacccagac catgagaatc aaggtggtcg agggcggccc tctccccttc
      961 gccttcgaca tcctggctac cagcttcatg tacggcagca aaaccttcat caaccacacc
     1021 cagggcatcc ccgacttctt taagcagtcc ttccctgagg gcttcacatg ggagagagtc
     1081 accacatacg aagacggggg cgtgctgacc gctacccagg acaccagcct ccaggacggc
     1141 tgcctcatct acaacgtcaa gatcagaggg gtgaacttcc catccaacgg ccctgtgatg
     1201 cagaagaaaa cactcggctg ggaggcctcc accgagatgc tgtaccccgc tgacggcggc
     1261 ctggaaggca gaagcgacat ggccctgaag ctcgtgggcg ggggccacct gatctgcaac
     1321 ttgaagacca catacagatc caagaaaccc gctaagaacc tcaagatgcc cggcgtctac
     1381 tatgtggaca gaagactgga aagaatcaag gaggccgaca aagagaccta cgtcgagcag
     1441 cacgaggtgg ctgtggccag atactgcgac ctccctagca aactggggca caaacttaat
     1501 tgattaatta atcaacatca gtctgataag ctatcaacat cagtctgata agctaaagct
     1561 tagctagtta cccagctttc ttgtacaaag tggtacgcgt gacggccgcg actctagatc
     1621 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     1681 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     1741 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     1801 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag
     1861 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     1921 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     1981 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     2041 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     2101 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     2161 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     2221 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     2281 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     2341 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     2401 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     2461 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     2521 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     2581 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     2641 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     2701 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     2761 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     2821 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     2881 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     2941 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     3001 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     3061 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     3121 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     3181 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     3241 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     3301 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3361 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     3421 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     3481 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     3541 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     3601 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     3661 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     3721 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     3781 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     3841 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     3901 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     3961 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4021 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4081 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4141 ttctgtggat aaccgtatta ccgcca
//
