LOCUS       sequence-178593-        4329 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway Mid-entry clone containing promoter sequence specific for 
            nrpb1b driving expression of eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4329)
  AUTHORS   Quillien A, Abdalla M, Yu J, Ou J, Zhu LJ, Lawson ND
  TITLE     Robust Identification of Developmentally Active Endothelial 
            Enhancers in Zebrafish Using FANS-Assisted ATAC-Seq.
  JOURNAL   Cell Rep. 2017 Jul 18;20(3):709-720. doi: 
            10.1016/j.celrep.2017.06.070.
  PUBMED    28723572
REFERENCE   2  (bases 1 to 4329)
  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.celrep.2017.06"; journalName: "Cell Rep"; date: 
            "2017-07-18- 18"; volume: "20"; issue: "3"; pages: "709-720"
COMMENT     Sequence Label: pMEnrp1bP1gfp
FEATURES             Location/Qualifiers
     source          1..4329
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      16..604
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     505..524
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     758..775
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(934..961)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(1053..1139)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     1188..1210
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1202..1219
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1203..1219
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1235..1334
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      2311..2320
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2317..2319,2320..2322,2323..3033)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2317..2319,2320..2322,2323..3033)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2362..2383)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2623..2642)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2970..2991
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3034..3099
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(3120..3219)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3236..3255)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3237..3255)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3260..3276)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3260..3276)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3378..3397)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3389..4198
                     /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(3456..3475)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
       61 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
      121 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
      181 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
      241 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
      301 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
      361 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
      421 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
      481 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
      541 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
      601 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
      661 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
      721 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
      781 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
      841 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatacgcg
      901 taccgctagc caggaagagt ttgtagaaac gcaaaaaggc catccgtcag gatggccttc
      961 tgcttagttt gatgcctggc agtttatggc gggcgtcctg cccgccaccc tccgggccgt
     1021 tgcttcacaa cgttcaaatc cgctcccggc ggatttgtcc tactcaggag agcgttcacc
     1081 gacaaacaac agataaaacg aaaggcccag tcttccgact gagcctttcg ttttatttga
     1141 tgcctggcag ttccctactc tcgcgttaac gctagcatgg atgttttccc agtcacgacg
     1201 ttgtaaaacg acggccagtc ttaagctcgg gccccaaata atgattttat tttgactgat
     1261 agtgacctgt tcgttgcaac aaattgatga gcaatgcttt tttataatgc caactttgta
     1321 caaaaaagca ggctcgctaa atgacatgtg tgatattaac tcaacagtga catgcttgct
     1381 ttcctgcacc tattaatgga ttaagactgc acatattttt tattttattt tactatgtga
     1441 attgggtaag cttaactgac agtagtgtat gtgtgtgaat gagagtgtat ggatgtttcc
     1501 caatgatggg ttgcagctgg aagaataatt attaatatta tttagatcag agcaaactat
     1561 ttttaactat tagaggacat accccttcct tcacccccat atatcttgtt ttgatgtttg
     1621 ccagggagga tcaagcatta cttagggggt cagacccccc cccctccccc cattcacatt
     1681 ctgacttaga gtaaaatatc taacttgtaa tgttagtgtt tactgaaacc ttatttgaaa
     1741 atgcagttat taacatatac aacagatgcg taaacaaaca tacctcaact tttttattga
     1801 cttcttctta ggttgtcacg ttaattgggg gtgccaaacc cctcatttct ctgcactgcg
     1861 ctcccacgca gaggagcgcg aagtctctct ggtttggccc ccggtctccg tgtctggttg
     1921 gcagagagag acagcgcgtc ggtctctgcc tggaaggggt gagagggttt gctcgtttcc
     1981 agtcctgccc attttctctc aaggaggagg aacgggagca accagtgctg ccgagagttt
     2041 tgacaccctc agtcttcata tcggatgctg acaccttcgc actagtgaga acctgctaaa
     2101 tatacccagc agtgcccgtg tacgcgcgac acagaggacg gaaatcagtc atatattagg
     2161 gagtgacccg actgctgaag cgcccgcaga cagcgagccg agcccaggac gcgctgtcaa
     2221 gcatctccaa cacttcggat tatgaaattg aagcaaacac agcagtggcg gtgactctac
     2281 gagaatctcc cgctttccag ggttgtgatc gccaccatgg tgagcaaggg cgaggagctg
     2341 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     2401 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     2461 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
     2521 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
     2581 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     2641 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     2701 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     2761 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     2821 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     2881 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     2941 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     3001 gggatcactc tcggcatgga cgagctgtac aagtccggac tcagatctcg agctcaagct
     3061 tcgaattctg cagtcgacgg taccgcgggc ccgggatcca ccggatctag ataactgata
     3121 cccagctttc ttgtacaaag ttggcattat aagaaagcat tgcttatcaa tttgttgcaa
     3181 cgaacaggtc actatcagtc aaaataaaat cattatttgc catccagctg atatccccta
     3241 tagtgagtcg tattacatgg tcatagctgt ttcctggcag ctctggcccg tgtctcaaaa
     3301 tctctgatgt tacattgcac aagataaaaa tatatcatca tgaacaataa aactgtctgc
     3361 ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa cgtcgaggcc
     3421 gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc gcgataatgt
     3481 cgggcaatca ggtgcgacaa tctatcgctt gtatgggaag cccgatgcgc cagagttgtt
     3541 tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg tcagactaaa
     3601 ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta ctcctgatga
     3661 tgcatggtta ctcaccactg cgatccccgg aaaaacagca ttccaggtat tagaagaata
     3721 tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc
     3781 gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca
     3841 atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc gtaatggctg
     3901 gcctgttgaa caagtctgga aagaaatgca taaacttttg ccattctcac cggattcagt
     3961 cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga aattaatagg
     4021 ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg ccatcctatg
     4081 gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa aatatggtat
     4141 tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt ttttctaatc
     4201 agaattggtt aattggttgt aacactggca gagcattacg ctgacttgac gggacggcgc
     4261 aagctcatga ccaaaatccc ttaacgtgag ttacgcgtcg ttccactgag cgtcagaccc
     4321 cgtagaaaa
//
