LOCUS       sequence-390726-        4341 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway entry vector for an inducible 3xFLAG-tagged TWIST2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4341)
  AUTHORS   Dutta AB, Lank DS, Przanowska RK, Przanowski P, Wang L, Nguyen B, 
            Walavalkar NM, Duarte FM, Guertin MJ
  TITLE     Kinetic networks identify TWIST2 as a key regulatory node in 
            adipogenesis.
  JOURNAL   Genome Res. 2023 Feb 21;33(3):314-31. doi: 10.1101/gr.277559.122.
  PUBMED    36810156
REFERENCE   2  (bases 1 to 4341)
  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: "10.1101/gr.277559";
            journalName: "Genome Res"; date: "2023-02-21- 21"; volume: "33"; 
            issue: "3"; pages: "314-31"
COMMENT     Sequence Label: pEN_TT 3xFLAG-TWIST2
FEATURES             Location/Qualifiers
     source          1..4341
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      120..206
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      298..325
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     333..356
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    375..474
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        506..820
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    514..532
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    550..568
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    585..603
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    621..639
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    657..675
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    692..710
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    728..746
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     793..817
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             829..894
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     protein_bind    complement(1419..1518)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1630..1649)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(1708..1727)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(2301..2834)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3023..3028,3029..3045,3046..3051))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      3708..4296
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4197..4216
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
       61 gctagcatgg atctcgggga cgtctaacta ctaagcgaga gtagggaact gccaggcatc
      121 aaataaaacg aaaggctcag tcggaagact gggcctttcg ttttatctgt tgtttgtcgg
      181 tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt gtgaagcaac
      241 ggcccggagg gtggcgggca ggacgcccgc cataaactgc caggcatcaa actaagcaga
      301 aggccatcct gacggatggc ctttttgcgt ttctacaaac tcttcctgtt agttagttac
      361 ttaagctcgg gccccaaata atgattttat tttgactgat agtgacctgt tcgttgcaac
      421 aaattgataa gcaatgcttt tttataatgc caactttgta caaaaaagca ggctttaaag
      481 gaaccaattc agtcgactgg atcccgagtt tactccctat cagtgataga gaacgtatgt
      541 cgagtttact ccctatcagt gatagagaac gatgtcgagt ttactcccta tcagtgatag
      601 agaacgtatg tcgagtttac tccctatcag tgatagagaa cgtatgtcga gtttactccc
      661 tatcagtgat agagaacgta tgtcgagttt atccctatca gtgatagaga acgtatgtcg
      721 agtttactcc ctatcagtga tagagaacgt atgtcgaggt aggcgtgtac ggtgggaggc
      781 ctatataagc agagctcgtt tagtgaaccg tcagatcgcc actagatgga ctacaaagac
      841 catgacggtg attataaaga tcatgacatc gattacaagg atgacgatga caagactagt
      901 atggaggagg gctccagctc gcccgtgtcc cccgtggaca gcctgggcac cagcgaggag
      961 gagctcgaga ggcagcccaa gcgcttcggc cggaagcggc gctacagcaa gaagtcgagc
     1021 gaagatggca gcccgacccc gggcaagcgc ggcaagaagg gcagccccag cgcgcagtcc
     1081 ttcgaggagc tgcagagcca gcgcatcctg gccaacgtgc gcgagcgcca gcgcacccag
     1141 tcgctcaacg aggccttcgc ggcgctgcgc aagatcatcc ccacgctgcc ctctgacaag
     1201 ctgagcaaga tccagacgct caagctggcc gccaggtaca tagacttcct ctaccaggtc
     1261 ctgcagagcg acgagatgga caataagatg accagctgca gctacgtggc ccacgagcgc
     1321 ctcagctacg ccttctccgt gtggcgcatg gagggcgcgt ggtccatgtc cgcctcccac
     1381 tagcaattgg aagactaatg cgttttcgag atatctagac ccagctttct tgtacaaagt
     1441 tggcattata agaaagcatt gcttatcaat ttgttgcaac gaacaggtca ctatcagtca
     1501 aaataaaatc attatttgcc atccagctgc agctctggcc cgtgtctcaa aatctctgat
     1561 gttacattgc acaagataaa aatatatcat catgaacaat aaaactgtct gcttacataa
     1621 acagtaatac aaggggtgtt atgagccata ttcaacggga aacgtcgagg ccgcgattaa
     1681 attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat
     1741 caggtgcgac aatctatcgc ttgtatggga agcccgatgc gccagagttg tttctgaaac
     1801 atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga
     1861 cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt
     1921 tactcaccac tgcgatcccc ggaaaaacag cattccaggt attagaagaa tatcctgatt
     1981 caggtgaaaa tattgttgat gcgctggcag tgtccctgcg ccggttgcat tcgattcctg
     2041 tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa
     2101 tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg
     2161 aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc
     2221 atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaggc cgggaagccg
     2281 atctcggctt gaacgaattg ttaggtggcg gtacttgggt cgatatcaaa gtgcatcact
     2341 tcttcccgta tgcccaactt tgtatagaga gccactgcgg gatcgtcacc gtaatctgct
     2401 tgcacgtaga tcacataagc accaagcgcg ttggcctcat gcttgaggag attgatgagc
     2461 gcggtggcaa tgccctgcct ccggtgctcg ccggagactg cgagatcata gatatagatc
     2521 tcactacgcg gctgctcaaa cttgggcaga acgtaagccg cgagagcgcc aacaaccgct
     2581 tcttggtcga aggcagcaag cgcgatgaat gtcttactac ggagcaagtt cccgaggtaa
     2641 tcggagtccg gctgatgttg ggagtaggtg gctacgtctc cgaactcacg accgaaaaga
     2701 tcaagagcag cccgcatgga tttgacttgg tcagggccga gcctacatgt gcgaatgatg
     2761 cccatacttg agccacctaa ctttgtttta gggcgactgc cctgctgcgt aacatcgttg
     2821 ctgctgcgta acatcgttgc tgctccataa catcaaacat cgacccacgg cgtaacgcgc
     2881 ttgctgcttg gatgcccgag gcatagactg tacaaaaaaa cagtcataac aagccatgaa
     2941 aaccgccact gcgccgttac caccgctgcg ttcggtcaag gttctggacc agttgcgtga
     3001 gcgcatacgc tacttgcatt acagtttacg aaccgaacag gcttatgtca actgggttcg
     3061 tgccttcatc cgtttccacg gtgtgcgtca cccggcaacc ttgggcagca gcgaagtcga
     3121 ggcatttctg tcctggctgg cgaacgagcg caaggtttcg gtctccacgc atcgtcaggc
     3181 attggcggcc ttgctgttct tctacggcaa ggtgctgtgc acggatctgc cctggcttca
     3241 ggagatcgga agacctcggc cgtcgcggcg cttgccggtg gtgctgaccc cggatgaagt
     3301 ggttcgcatc ctcggttttc tggaaggcga gcatcgtttg ttcgcccagg actctagcta
     3361 tagttctagt ggttggctac taataggttg tattgatgtt ggacgagtcg gaatcgcaga
     3421 ccgataccag gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca
     3481 gaaacggctt tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca
     3541 tttgatgctc gatgagtttt tctaatcaga attggttaat tggttgtaac actggcagag
     3601 cattacgctg acttgacggg acggcgcaag ctcatgacca aaatccctta acgtgagttt
     3661 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     3721 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     3781 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     3841 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     3901 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     3961 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     4021 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     4081 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     4141 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     4201 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     4261 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     4321 cggttcctgg ccttttgctg g
//
