LOCUS       sequence-272002-        4280 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway compatible middle entry clone contains Firefly Luciferase2 
            insert driven by minimal promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4280)
  AUTHORS   Sissaoui S, Yu J, Yan A, Li R, Zhu LJ, Kucukural A, Yukselen O, 
            Lawson ND
  TITLE     Genomic Characterization of Endothelial Enhancers Reveals a 
            Multifunctional Role for NR2F2 in Regulation of Arteriovenous Gene 
            Expression.
  JOURNAL   Circ Res. 2020 Feb 17. doi: 10.1161/CIRCRESAHA.119.316075.
  PUBMED    32065070
REFERENCE   2  (bases 1 to 4280)
  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"; journalName: "Circ Res. 
            2020 Feb 17. doi: 10.1161/CIRCRESAHA.119.316075."
COMMENT     Sequence Label: pME-minP:luc2
FEATURES             Location/Qualifiers
     source          1..4280
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     98..115
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(274..301)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(393..479)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     528..550
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     542..559
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     543..559
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    575..674
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        681..712
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      739..748
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             745..2397
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     protein_bind    complement(2411..2510)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2527..2546)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2528..2546)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2551..2567)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2551..2567)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2669..2688)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2680..3489
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2747..2766)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3636..4224
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4125..4144
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
       61 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
      121 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
      181 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatacgcg
      241 taccgctagc caggaagagt ttgtagaaac gcaaaaaggc catccgtcag gatggccttc
      301 tgcttagttt gatgcctggc agtttatggc gggcgtcctg cccgccaccc tccgggccgt
      361 tgcttcacaa cgttcaaatc cgctcccggc ggatttgtcc tactcaggag agcgttcacc
      421 gacaaacaac agataaaacg aaaggcccag tcttccgact gagcctttcg ttttatttga
      481 tgcctggcag ttccctactc tcgcgttaac gctagcatgg atgttttccc agtcacgacg
      541 ttgtaaaacg acggccagtc ttaagctcgg gccccaaata atgattttat tttgactgat
      601 agtgacctgt tcgttgcaac aaattgatga gcaatgcttt tttataatgc caactttgta
      661 caaaaaagca ggctagacac tagagggtat ataatggaag ctcgacttcc agcttggcaa
      721 tccggtactg ttggtaaagc caccatggaa gatgccaaaa acattaagaa gggcccagcg
      781 ccattctacc cactcgaaga cgggaccgcc ggcgagcagc tgcacaaagc catgaagcgc
      841 tacgccctgg tgcccggcac catcgccttt accgacgcac atatcgaggt ggacattacc
      901 tacgccgagt acttcgagat gagcgttcgg ctggcagaag ctatgaagcg ctatgggctg
      961 aatacaaacc atcggatcgt ggtgtgcagc gagaatagct tgcagttctt catgcccgtg
     1021 ttgggtgccc tgttcatcgg tgtggctgtg gccccagcta acgacatcta caacgagcgc
     1081 gagctgctga acagcatggg catcagccag cccaccgtcg tattcgtgag caagaaaggg
     1141 ctgcaaaaga tcctcaacgt gcaaaagaag ctaccgatca tacaaaagat catcatcatg
     1201 gatagcaaga ccgactacca gggcttccaa agcatgtaca ccttcgtgac ttcccatttg
     1261 ccacccggct tcaacgagta cgacttcgtg cccgagagct tcgaccggga caaaaccatc
     1321 gccctgatca tgaacagtag tggcagtacc ggattgccca agggcgtagc cctaccgcac
     1381 cgcaccgctt gtgtccgatt cagtcatgcc cgcgacccca tcttcggcaa ccagatcatc
     1441 cccgacaccg ctatcctcag cgtggtgcca tttcaccacg gcttcggcat gttcaccacg
     1501 ctgggctact tgatctgcgg ctttcgggtc gtgctcatgt accgcttcga ggaggagcta
     1561 ttcttgcgca gcttgcaaga ctataagatt caatctgccc tgctggtgcc cacactattt
     1621 agcttcttcg ctaagagcac tctcatcgac aagtacgacc taagcaactt gcacgagatc
     1681 gccagcggcg gggcgccgct cagcaaggag gtaggtgagg ccgtggccaa acgcttccac
     1741 ctaccaggca tccgccaggg ctacggcctg acagaaacaa ccagcgccat tctgatcacc
     1801 cccgaagggg acgacaagcc tggcgcagta ggcaaggtgg tgcccttctt cgaggctaag
     1861 gtggtggact tggacaccgg taagacactg ggtgtgaacc agcgcggcga gctgtgcgtc
     1921 cgtggcccca tgatcatgag cggctacgtt aacaaccccg aggctacaaa cgctctcatc
     1981 gacaaggacg gctggctgca cagcggcgac atcgcctact gggacgagga cgagcacttc
     2041 ttcatcgtgg accggctgaa gagcctgatc aaatacaagg gctaccaggt agccccagcc
     2101 gaactggaga gcatcctgct gcaacacccc aacatcttcg acgccggggt cgccggcctg
     2161 cccgacgacg atgccggcga gctgcccgcc gcagtcgtcg tgctggaaca cggtaaaacc
     2221 atgaccgaga aggagatcgt ggactatgtg gccagccagg ttacaaccgc caagaagctg
     2281 cgcggtggtg ttgtgttcgt ggacgaggtg cctaaaggac tgaccggcaa gttggacgcc
     2341 cgcaagatcc gcgagattct cattaaggcc aagaagggcg gcaagatcgc cgtgtaataa
     2401 ttctagagtc acccagcttt cttgtacaaa gttggcatta taagaaagca ttgcttatca
     2461 atttgttgca acgaacaggt cactatcagt caaaataaaa tcattatttg ccatccagct
     2521 gatatcccct atagtgagtc gtattacatg gtcatagctg tttcctggca gctctggccc
     2581 gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc atgaacaata
     2641 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa
     2701 acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct
     2761 cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg
     2821 ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg
     2881 gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt
     2941 actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta
     3001 ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc
     3061 cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc
     3121 gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag
     3181 cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca
     3241 ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg
     3301 aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt
     3361 gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa
     3421 aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag
     3481 tttttctaat cagaattggt taattggttg taacactggc agagcattac gctgacttga
     3541 cgggacggcg caagctcatg accaaaatcc cttaacgtga gttacgcgtc gttccactga
     3601 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     3661 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     3721 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     3781 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     3841 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     3901 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     3961 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     4021 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     4081 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     4141 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     4201 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     4261 ttttgctggc cttttgctca
//
