LOCUS       62056_20385        4139 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pT-GTW6-CMV-mKate_2xmiR221ts3'UTR, complete sequence.
ACCESSION   MT891365
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4139)
  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 4139)
  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 4139)
  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..4139
                     /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"
     CDS             748..765
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             766..1473
                     /label=PAmKate
                     /note="monomeric photoactivatable red fluorescent protein 
                     (Gunewardene et al., 2011)"
     misc_feature    1485..1528
                     /label=2xmiR221-TS
                     /note="2xmiR221-TS"
     protein_bind    complement(1543..1567)
                     /label=attB2
                     /note="recombination site for the Gateway(R) BP reaction"
     polyA_signal    1696..1817
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(1824..2279)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        2306..2410
                     /label=AmpR promoter
     CDS             2411..3268
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      3442..4030
                     /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 tggatcccgc caccatgcac catcaccatc accatgtgtc taagggcgaa
      781 gagctgatta aggagaacat gcacatgaag ctgtacatgg agggcaccgt gaacaaccac
      841 cacttcaagt gcacatccga gggcgaaggc aagccctacg agggcaccca gaccatgaga
      901 atcaaggtgg tcgagggcgg ccctctcccc ttcgccttcg acatcctggc taccagcttc
      961 atgtacggca gcaaaacctt catcaaccac acccagggca tccccgactt ctttaagcag
     1021 tccttccctg agggcttcac atgggagaga gtcaccacat acgaagacgg gggcgtgctg
     1081 accgctaccc aggacaccag cctccaggac ggctgcctca tctacaacgt caagatcaga
     1141 ggggtgaact tcccatccaa cggccctgtg atgcagaaga aaacactcgg ctgggaggcc
     1201 tccaccgaga tgctgtaccc cgctgacggc ggcctggaag gcagaagcga catggccctg
     1261 aagctcgtgg gcgggggcca cctgatctgc aacttgaaga ccacatacag atccaagaaa
     1321 cccgctaaga acctcaagat gcccggcgtc tactatgtgg acagaagact ggaaagaatc
     1381 aaggaggccg acaaagagac ctacgtcgag cagcacgagg tggctgtggc cagatactgc
     1441 gacctcccta gcaaactggg gcacaaactt aattgattaa ttaaaaatct acattgtatg
     1501 ccaggtaaat ctacattgta tgccaggtaa gcttagctag ttacccagct ttcttgtaca
     1561 aagtggtacg cgtgacggcc gcgactctag atcataatca gccataccac atttgtagag
     1621 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     1681 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     1741 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     1801 ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt
     1861 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     1921 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     1981 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     2041 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     2101 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     2161 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     2221 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     2281 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     2341 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     2401 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     2461 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     2521 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     2581 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     2641 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     2701 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     2761 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     2821 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     2881 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     2941 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     3001 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     3061 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     3121 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     3181 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     3241 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     3301 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     3361 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     3421 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     3481 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     3541 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     3601 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     3661 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     3721 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     3781 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     3841 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     3901 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     3961 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     4021 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     4081 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcca
//
