LOCUS       62056_11790        4700 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pGLM91(P_TRE-DsRed(FF4)), complete sequence.
ACCESSION   MT891335
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4700)
  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 4700)
  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 4700)
  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..4700
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      1..266
                     /note="misc_signal; INS"
                     /regulatory_class="other"
     polyA_signal    283..507
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     misc_feature    529..553
                     /label=bidirectional promoter artefact
                     /note="bidirectional promoter artefact"
     promoter        554..868
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     misc_feature    896..905
                     /label=Kozak sequence
                     /note="Kozak sequence"
     ncRNA           1181..1195
                     /label=pre_miRNA
                     /note="miR-30a loop"
                     /ncRNA_class="pre_miRNA"
     ncRNA           1198..1219
                     /label=miRNA
                     /note="miR-FF4"
                     /ncRNA_class="miRNA"
     ncRNA           1223..1351
                     /label=3' miR-30a
                     /note="sequence downstream of the 71-nt precursor of the
                     human miR-30a microRNA (Zeng et al., 2002)"
     misc_binding    1988..2009
                     /label=miR-FF4 binding site
                     /bound_moiety="miR-FF4"
                     /note="mir FF4 target"
     misc_binding    2010..2031
                     /label=miR-FF4 binding site
                     /bound_moiety="miR-FF4"
                     /note="mir FF4 target"
     misc_binding    2032..2053
                     /label=miR-FF4 binding site
                     /bound_moiety="miR-FF4"
                     /note="mir FF4 target"
     promoter        complement(2091..2109)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     polyA_signal    2267..2388
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     regulatory      2478..2767
                     /note="misc_signal; INS"
                     /regulatory_class="other"
     rep_origin      complement(2855..3443)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3617..4474)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(4475..4579)
                     /label=AmpR promoter
ORIGIN
        1 gtcttctcca tcggatcctg atcaggggag ctcacgggga cagccccccc ccaaagcccc
       61 cagggatgta attacgtccc tcccccgcta gggggcagca gcgaccgccc ggggctccgc
      121 tccggtccgg cgctcccccc gcatcccgag ccggcagcgt gcggggacag cccgggcacg
      181 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
      241 acacctgggg gatacgggga aaaccgcggc atatggcctc gactgtgcct tctagttgcc
      301 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
      361 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
      421 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
      481 atgctgggga tgcggtgggc tctatggctt ctgagggcta gcgatatccc gtacacgcca
      541 cctcgacata ctcgagttta ctccctatca gtgatagaga acgtatgtcg agtttactcc
      601 ctatcagtga tagagaacga tgtcgagttt actccctatc agtgatagag aacgtatgtc
      661 gagtttactc cctatcagtg atagagaacg tatgtcgagt ttactcccta tcagtgatag
      721 agaacgtatg tcgagtttat ccctatcagt gatagagaac gtatgtcgag tttactccct
      781 atcagtgata gagaacgtat gtcgaggtag gcgtgtacgg tgggaggcct atataagcag
      841 agctcgttta gtgaaccgtc agatcgcctg gagatttcga gctcggtgcg gccgcgccac
      901 catggcctcc tccgaggacg tcatcaagga gttcatgcgc ttcaaggtgc gcatggaggg
      961 ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gaggtgagta tgtgctcgct
     1021 tcggcagcac atatactatg tcgaatgagg cttcagtact ttacagaatc gttgcctgca
     1081 catcttggaa acacttgctg ggattacttc ttcaggttaa cccaacagaa ggtacgtatg
     1141 ctgttgacag tgagcgccgc ttgaagtttt taattaaata gtgaagccac agatgtattt
     1201 aattaaagac ttcaagcggt gcctactgcc tcggagaatt caaggggcta ctttaggagc
     1261 aattatcttg tttactaaaa ctgaatacct tgctatctct ttgatacatt tttacaaagc
     1321 tgaattaaaa tggtataaat taaatcactt ttttcaattg tttccttttt tttcctcagg
     1381 gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggc ggccccctgc
     1441 ccttcgcctg ggacatcctg tccccccagt tccagtacgg ctccaaggtg tacgtgaagc
     1501 accccgccga catccccgac tacaagaagc tgtccttccc cgagggcttc aagtgggagc
     1561 gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg
     1621 acggctcctt catctacaag gtgaagttca tcggcgtgaa cttcccctcc gacggccccg
     1681 taatgcagaa gaagactatg ggctgggagg cctccaccga gcgcctgtac ccccgcgacg
     1741 gcgtgctgaa gggcgagatc cacaaggccc tgaagctgaa ggacggcggc cactacctgg
     1801 tggagttcaa gtccatctac atggccaaga agcccgtgca gctgcccggc tactactacg
     1861 tggactccaa gctggacatc acctcccaca acgaggacta caccatcgtg gagcagtacg
     1921 agcgcgccga gggccgccac cacctgttcc tgagatctcg agctcaagct tactaacatg
     1981 cttcgaaccg cttgaagtct ttaattaaac cgcttgaagt ctttaattaa accgcttgaa
     2041 gtctttaatt aaaggcgcgc ctgtacaagt agtaatctag agggccctat tctatagtgt
     2101 cacctaaatg ctagagctcg ctgatcagtc cgtcgacgga tccaccggat ctagataact
     2161 gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca
     2221 cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc
     2281 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     2341 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaac gcgtaaattg
     2401 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     2461 accaataggc cgaaatcggg gagctcacgg ggacagcccc cccccaaagc ccccagggat
     2521 gtaattacgt ccctcccccg ctagggggca gcagcgaccg cccggggctc cgctccggtc
     2581 cggcgctccc cccgcatccc gagccggcag cgtgcgggga cagcccgggc acggggaagg
     2641 tggcacggga tcgctttcct ctgaacgctt ctcgctgctc tttgagcctg cagacacctg
     2701 ggggatacgg ggaaaatgat caagatcttc tacactggga cgcaaggatt atcctcgagg
     2761 cgcgcgaaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     2821 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     2881 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     2941 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     3001 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     3061 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     3121 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     3181 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     3241 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     3301 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     3361 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     3421 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     3481 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     3541 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     3601 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     3661 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     3721 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     3781 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     3841 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     3901 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     3961 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     4021 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     4081 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     4141 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     4201 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     4261 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     4321 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     4381 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     4441 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     4501 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     4561 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg
     4621 gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     4681 aagccagtat ctgctccctg
//
