LOCUS       sequence-272000-        4222 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway compatible middle entry clone contains Firefly Luciferase2 
            without minimal promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4222)
  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 4222)
  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-luc2
FEATURES             Location/Qualifiers
     source          1..4222
                     /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"
     regulatory      681..690
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             687..2339
                     /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(2353..2452)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2469..2488)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2470..2488)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2493..2509)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2493..2509)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2611..2630)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2622..3431
                     /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(2689..2708)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3578..4166
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4067..4086
                     /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 ggctggtaaa gccaccatgg aagatgccaa aaacattaag aagggcccag
      721 cgccattcta cccactcgaa gacgggaccg ccggcgagca gctgcacaaa gccatgaagc
      781 gctacgccct ggtgcccggc accatcgcct ttaccgacgc acatatcgag gtggacatta
      841 cctacgccga gtacttcgag atgagcgttc ggctggcaga agctatgaag cgctatgggc
      901 tgaatacaaa ccatcggatc gtggtgtgca gcgagaatag cttgcagttc ttcatgcccg
      961 tgttgggtgc cctgttcatc ggtgtggctg tggccccagc taacgacatc tacaacgagc
     1021 gcgagctgct gaacagcatg ggcatcagcc agcccaccgt cgtattcgtg agcaagaaag
     1081 ggctgcaaaa gatcctcaac gtgcaaaaga agctaccgat catacaaaag atcatcatca
     1141 tggatagcaa gaccgactac cagggcttcc aaagcatgta caccttcgtg acttcccatt
     1201 tgccacccgg cttcaacgag tacgacttcg tgcccgagag cttcgaccgg gacaaaacca
     1261 tcgccctgat catgaacagt agtggcagta ccggattgcc caagggcgta gccctaccgc
     1321 accgcaccgc ttgtgtccga ttcagtcatg cccgcgaccc catcttcggc aaccagatca
     1381 tccccgacac cgctatcctc agcgtggtgc catttcacca cggcttcggc atgttcacca
     1441 cgctgggcta cttgatctgc ggctttcggg tcgtgctcat gtaccgcttc gaggaggagc
     1501 tattcttgcg cagcttgcaa gactataaga ttcaatctgc cctgctggtg cccacactat
     1561 ttagcttctt cgctaagagc actctcatcg acaagtacga cctaagcaac ttgcacgaga
     1621 tcgccagcgg cggggcgccg ctcagcaagg aggtaggtga ggccgtggcc aaacgcttcc
     1681 acctaccagg catccgccag ggctacggcc tgacagaaac aaccagcgcc attctgatca
     1741 cccccgaagg ggacgacaag cctggcgcag taggcaaggt ggtgcccttc ttcgaggcta
     1801 aggtggtgga cttggacacc ggtaagacac tgggtgtgaa ccagcgcggc gagctgtgcg
     1861 tccgtggccc catgatcatg agcggctacg ttaacaaccc cgaggctaca aacgctctca
     1921 tcgacaagga cggctggctg cacagcggcg acatcgccta ctgggacgag gacgagcact
     1981 tcttcatcgt ggaccggctg aagagcctga tcaaatacaa gggctaccag gtagccccag
     2041 ccgaactgga gagcatcctg ctgcaacacc ccaacatctt cgacgccggg gtcgccggcc
     2101 tgcccgacga cgatgccggc gagctgcccg ccgcagtcgt cgtgctggaa cacggtaaaa
     2161 ccatgaccga gaaggagatc gtggactatg tggccagcca ggttacaacc gccaagaagc
     2221 tgcgcggtgg tgttgtgttc gtggacgagg tgcctaaagg actgaccggc aagttggacg
     2281 cccgcaagat ccgcgagatt ctcattaagg ccaagaaggg cggcaagatc gccgtgtaat
     2341 aattctagag tcacccagct ttcttgtaca aagttggcat tataagaaag cattgcttat
     2401 caatttgttg caacgaacag gtcactatca gtcaaaataa aatcattatt tgccatccag
     2461 ctgatatccc ctatagtgag tcgtattaca tggtcatagc tgtttcctgg cagctctggc
     2521 ccgtgtctca aaatctctga tgttacattg cacaagataa aaatatatca tcatgaacaa
     2581 taaaactgtc tgcttacata aacagtaata caaggggtgt tatgagccat attcaacggg
     2641 aaacgtcgag gccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg
     2701 ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg cttgtatggg aagcccgatg
     2761 cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga
     2821 tggtcagact aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc
     2881 gtactcctga tgatgcatgg ttactcacca ctgcgatccc cggaaaaaca gcattccagg
     2941 tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgttcctgc
     3001 gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc
     3061 tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg
     3121 agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt ttgccattct
     3181 caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg
     3241 ggaaattaat aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc
     3301 ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc
     3361 aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg
     3421 agtttttcta atcagaattg gttaattggt tgtaacactg gcagagcatt acgctgactt
     3481 gacgggacgg cgcaagctca tgaccaaaat cccttaacgt gagttacgcg tcgttccact
     3541 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3601 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3661 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3721 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3781 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     3841 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     3901 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     3961 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4021 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4081 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4141 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     4201 ccttttgctg gccttttgct ca
//
