LOCUS       sequence-183531-        5500 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway entry vector for inducible expression of Luciferase with 
            N-terminal 3xFLAG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5500)
  AUTHORS   Bajikar SS, Wang CC, Borten MA, Pereira EJ, Atkins KA, Janes KA
  TITLE     Tumor-Suppressor Inactivation of GDF11 Occurs by Precursor 
            Sequestration in Triple-Negative Breast Cancer.
  JOURNAL   Dev Cell. 2017 Nov 20;43(4):418-435.e13. doi: 
            10.1016/j.devcel.2017.10.027.
  PUBMED    29161592
REFERENCE   2  (bases 1 to 5500)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.devcel.2017.10"; journalName: "Dev Cell"; date: 
            "2017-11-20- 20"; volume: "43"; issue: "4"; pages: "418-435.e13"
COMMENT     Sequence Label: pEN_TT 3xFLAG Luciferase
FEATURES             Location/Qualifiers
     source          1..5500
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      25..52
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     60..83
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    102..201
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        233..547
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    241..259
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    277..295
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    312..330
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    348..366
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    384..402
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    419..437
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    455..473
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     520..544
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             557..622
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(689..707)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     protein_bind    complement(2305..2404)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2516..2535)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(2594..2613)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(3187..3720)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3909..3914,3915..3931,3932..3937))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      4594..5182
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5083..5102
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      5347..5433
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 aaactgccag gcatcaaact aagcagaagg ccatcctgac ggatggcctt tttgcgtttc
       61 tacaaactct tcctgttagt tagttactta agctcgggcc ccaaataatg attttatttt
      121 gactgatagt gacctgttcg ttgcaacaaa ttgataagca atgctttttt ataatgccaa
      181 ctttgtacaa aaaagcaggc tttaaaggaa ccaattcagt cgactggatc ccgagtttac
      241 tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgat
      301 gtcgagttta ctccctatca gtgatagaga acgtatgtcg agtttactcc ctatcagtga
      361 tagagaacgt atgtcgagtt tactccctat cagtgataga gaacgtatgt cgagtttatc
      421 cctatcagtg atagagaacg tatgtcgagt ttactcccta tcagtgatag agaacgtatg
      481 tcgaggtagg cgtgtacggt gggaggccta tataagcaga gctcgtttag tgaaccgtca
      541 gatcgccact agtatggact acaaagacca tgacggtgat tataaagatc atgacatcga
      601 ttacaaggat gacgatgaca agatggaaga cgccaaaaac ataaagaaag gcccggcgcc
      661 attctatccg ctggaagatg gaaccgctgg agagcaactg cataaggcta tgaagagata
      721 cgccctggtt cctggaacaa ttgcttttac agatgcacat atcgaggtgg acatcactta
      781 cgctgagtac ttcgaaatgt ccgttcggtt ggcagaagct atgaaacgat atgggctgaa
      841 tacaaatcac agaatcgtcg tatgcagtga aaactctctt caattcttta tgccggtgtt
      901 gggcgcgtta tttatcggag ttgcagttgc gcccgcgaac gacatttata atgaacgtga
      961 attgctcaac agtatgggca tttcgcagcc taccgtggtg ttcgtttcca aaaaggggtt
     1021 gcaaaaaatt ttgaacgtgc aaaaaaagct cccaatcatc caaaaaatta ttatcatgga
     1081 ttctaaaacg gattaccagg gatttcagtc gatgtacacg ttcgtcacat ctcatctacc
     1141 tcccggtttt aatgaatacg attttgtgcc agagtccttc gatagggaca agacaattgc
     1201 actgatcatg aactcctctg gatctactgg tctgcctaaa ggtgtcgctc tgcctcatag
     1261 aactgcctgc gtgagattct cgcatgccag agatcctatt tttggcaatc aaatcattcc
     1321 ggatactgcg attttaagtg ttgttccatt ccatcacggt tttggaatgt ttactacact
     1381 cggatatttg atatgtggat ttcgagtcgt cttaatgtat agatttgaag aagagctgtt
     1441 tctgaggagc cttcaggatt acaagattca aagtgcgctg ctggtgccaa ccctattctc
     1501 cttcttcgcc aaaagcactc tgattgacaa atacgattta tctaatttac acgaaattgc
     1561 ttctggtggc gctcccctct ctaaggaagt cggggaagcg gttgccaaga ggttccatct
     1621 gccaggtatc aggcaaggat atgggctcac tgagactaca tcagctattc tgattacacc
     1681 cgagggggat gataaaccgg gcgcggtcgg taaagttgtt ccattttttg aagcgaaggt
     1741 tgtggatctg gataccggga aaacgctggg cgttaatcaa agaggcgaac tgtgtgtgag
     1801 aggtcctatg attatgtccg gttatgtaaa caatccggaa gcgaccaacg ccttgattga
     1861 caaggatgga tggctacatt ctggagacat agcttactgg gacgaagacg aacacttctt
     1921 catcgttgac cgcctgaagt ctctgattaa gtacaaaggc tatcaggtgg ctcccgctga
     1981 attggaatcc atcttgctcc aacaccccaa catcttcgac gcaggtgtcg caggtcttcc
     2041 cgacgatgac gccggtgaac ttcccgccgc cgttgttgtt ttggagcacg gaaagacgat
     2101 gacggaaaaa gagatcgtgg attacgtcgc cagtcaagta acaaccgcga aaaagttgcg
     2161 cggaggagtt gtgtttgtgg acgaagtacc gaaaggtctt accggaaaac tcgacgcaag
     2221 aaaaatcaga gagatcctca taaaggccaa gaagggcgga aagatctgag aattggaaga
     2281 ctaatgcgtt ttcgagatat ctagacccag ctttcttgta caaagttggc attataagaa
     2341 agcattgctt atcaatttgt tgcaacgaac aggtcactat cagtcaaaat aaaatcatta
     2401 tttgccatcc agctgcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa
     2461 gataaaaata tatcatcatg aacaataaaa ctgtctgctt acataaacag taatacaagg
     2521 ggtgttatga gccatattca acgggaaacg tcgaggccgc gattaaattc caacatggat
     2581 gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc
     2641 tatcgcttgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc
     2701 gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct
     2761 cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg
     2821 atccccggaa aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt
     2881 gttgatgcgc tggcagtgtc cctgcgccgg ttgcattcga ttcctgtttg taattgtcct
     2941 tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg
     3001 gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa
     3061 gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca
     3121 cttgataacc ttatttttga cgaggggaaa ttaggccggg aagccgatct cggcttgaac
     3181 gaattgttag gtggcggtac ttgggtcgat atcaaagtgc atcacttctt cccgtatgcc
     3241 caactttgta tagagagcca ctgcgggatc gtcaccgtaa tctgcttgca cgtagatcac
     3301 ataagcacca agcgcgttgg cctcatgctt gaggagattg atgagcgcgg tggcaatgcc
     3361 ctgcctccgg tgctcgccgg agactgcgag atcatagata tagatctcac tacgcggctg
     3421 ctcaaacttg ggcagaacgt aagccgcgag agcgccaaca accgcttctt ggtcgaaggc
     3481 agcaagcgcg atgaatgtct tactacggag caagttcccg aggtaatcgg agtccggctg
     3541 atgttgggag taggtggcta cgtctccgaa ctcacgaccg aaaagatcaa gagcagcccg
     3601 catggatttg acttggtcag ggccgagcct acatgtgcga atgatgccca tacttgagcc
     3661 acctaacttt gttttagggc gactgccctg ctgcgtaaca tcgttgctgc tgcgtaacat
     3721 cgttgctgct ccataacatc aaacatcgac ccacggcgta acgcgcttgc tgcttggatg
     3781 cccgaggcat agactgtaca aaaaaacagt cataacaagc catgaaaacc gccactgcgc
     3841 cgttaccacc gctgcgttcg gtcaaggttc tggaccagtt gcgtgagcgc atacgctact
     3901 tgcattacag tttacgaacc gaacaggctt atgtcaactg ggttcgtgcc ttcatccgtt
     3961 tccacggtgt gcgtcacccg gcaaccttgg gcagcagcga agtcgaggca tttctgtcct
     4021 ggctggcgaa cgagcgcaag gtttcggtct ccacgcatcg tcaggcattg gcggccttgc
     4081 tgttcttcta cggcaaggtg ctgtgcacgg atctgccctg gcttcaggag atcggaagac
     4141 ctcggccgtc gcggcgcttg ccggtggtgc tgaccccgga tgaagtggtt cgcatcctcg
     4201 gttttctgga aggcgagcat cgtttgttcg cccaggactc tagctatagt tctagtggtt
     4261 ggctactaat aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc
     4321 ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc
     4381 aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg
     4441 agtttttcta atcagaattg gttaattggt tgtaacactg gcagagcatt acgctgactt
     4501 gacgggacgg cgcaagctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     4561 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     4621 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     4681 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     4741 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     4801 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     4861 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     4921 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     4981 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     5041 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     5101 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     5161 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5221 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     5281 tattaccgct agcatggatc tcggggacgt ctaactacta agcgagagta gggaactgcc
     5341 aggcatcaaa taaaacgaaa ggctcagtcg gaagactggg cctttcgttt tatctgttgt
     5401 ttgtcggtga acgctctcct gagtaggaca aatccgccgg gagcggattt gaacgttgtg
     5461 aagcaacggc ccggagggtg gcgggcagga cgcccgccat
//
