LOCUS       sequence-178748-        7663 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  contains dll4 enhancer 1 sequence and a basal promoter driving 
            expression of eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7663)
  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 7663)
  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: pT2-cryR;dll4CE1-basEgfp
FEATURES             Location/Qualifiers
     source          1..7663
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     646..662
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    682..702
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    1356..1380
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1457..1466
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1463..1465,1466..1468,1469..2179)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1463..1465,1466..1468,1469..2179)
                     /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(1508..1529)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1769..1788)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2116..2137
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2180..2245
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2369..2490
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(2497..2521)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2547..2567)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     complement(2575..2591)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    2607..2741
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(3180..3367,3368..3379)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(3413..3429)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3413..3429)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3466..3486)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3466..3484)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3505..3521)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3518..3540)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3529..3545
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3553..3559,3560..3577,3578..3583))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3598..3619
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3736..3753)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3907..4495)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3987..4006)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4666..5457,5458..5526))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5289..5308
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5527..5631)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5794..5815)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(6164..6175,6176..6338)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    6707..6828
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(6951..7661)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEDDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGHHSTGGMDELYK"
     primer_bind     complement(7031..7054)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(7229..7248)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     7498..7516
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      join(7658..7663,1..4)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 tggcgaccgg taatgtcaga cctggtaact ccttactgta acttattgca cctgatgaac
       61 aaaagatgtg aactgaacgc aagtaggagg ctaatagagg ctagtacagt acaaagatcc
      121 ccaaatgatg tgtgccggcc ttagtggagg cagacccctg gttatatacc tgtgctgcag
      181 aaagaggggc ttacaaatgg atctctctgg aatggcatta tctcatggtg ggaacagtgt
      241 tgtcagcaga atcagcgggg tggactctga cctgagaatg acagtctagc actgaatggc
      301 tcagacaata ctcttgtggc caactatcac cggaaaataa gtgcctgcca aaacaaacca
      361 gaccagatga agctacatgg ctgtagaacg tcagtattta atgtgttgtg gccacttggt
      421 tcaccagatt taagagctaa tttttttaaa tatcacagat aatcaatcgc tttttttttt
      481 cttgactgaa aagcttagac tggaaaattc tttgaaaaag tttgcactac agaaaataaa
      541 tacataacat cctgcactga atgcacacta ttaatagaaa agagagcatt attgtaggaa
      601 atgtattgta tatgtacatt ctgtgcattt cacaattttc tcgagcagga aacagctatg
      661 accatgatta cgccaagcta tcaactttgt atagaaaagt tgcagctgaa cactgaaaag
      721 gcagagaaaa ggctgccctc aggaacagcc gaccgttttt catgataagg atatttcatt
      781 gtttttgcaa caagacagct tgagtttaaa aaaaaaaaaa aaagtcctac agccgagcgg
      841 ctcagttgtt tggcggtgat ataatgcccg ttcagtcttt gttgtgtggg tgtcagttgg
      901 cggttgttga gagggggggc atgagtagtg ttgccccttt gtttgttttc aatggctcat
      961 aaacagtctg cataaaccga tagaaaaagg ccagcttccc cttcgctctg tgtgcggctg
     1021 attcctcgtt cacactcctc gggtcctacg actctgggat tgttttgagc tgtccagcga
     1081 ggaaggaagc cagtgtgaga aagtggctgt ttttttggag tggacaggat ttggcctcat
     1141 cgcgcgcggt tcctcttttc gtcggctgaa acaatgcagt ggcgtcacac ccctcgactg
     1201 gtgctcggcg tttcacatgg aaaaatgttc ccagaattcc gctcccaggc tcagaggtaa
     1261 ggctgggaat ggagagcgga ggaaacgcag agcgagagac gaacaggaat ggaggaaaag
     1321 ccagagacct gagcagaact gaacagtgct catatgcaag tttgtacaaa aaagcaggct
     1381 agactctaga gggtatataa tggatctcgc gtctcagcct cactttgagc tcctccacac
     1441 gctagcgcta ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt
     1501 gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga
     1561 gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa
     1621 gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag
     1681 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     1741 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     1801 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     1861 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     1921 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     1981 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     2041 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
     2101 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     2161 catggacgag ctgtacaagt ccggactcag atctcgagct caagcttcga attctgcagt
     2221 cgacggtacc gcgggcccgg gatccaccgg atctagataa ctgatcataa tcagccatac
     2281 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     2341 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     2401 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     2461 tggtttgtcc aaactcatca atgtatctta acgcgtaccc agctttcttg tacaaagtgg
     2521 ggcgcgccta cgtaactagt ctcgagcaac tttattatac atagttgata attcactggc
     2581 cgtcgtttta cggtaccatc gatgatgatc cagacatgat aagatacatt gatgagtttg
     2641 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     2701 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     2761 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct
     2821 acaaatgtgg tatggctgat tatgatcctc tagatcagat ctgcgaagat acggccacgg
     2881 gtgctcttga tcctgtggct gattttggac tgtgctgctc gcagctgctg atgaatcaca
     2941 tacttcctcc attttcttcc actgattgac tgttataatt tccctaattt ccaggtcaag
     3001 gtgctgtgca ttgtggtaat agatgtgaca tgacgtcact tccaaaggac caatgaacat
     3061 gtctgaccaa tttcatataa tgtgaaaacg attttcatag gcagaataaa taacatttaa
     3121 attaaactgg gcatcagcgc aattcaattg gtttggtaat agcaagggaa aatagaatga
     3181 agtgatctcc aaaaaataag tactttttga ctgtaaataa aattgtaagg agtaaaaagt
     3241 actttttttt ctaaaaaaat gtaattaagt aaaagtaaaa gtattgattt ttaattgtac
     3301 tcaagtaaag taaaaatccc caaaaataat acttaagtac agtaatcaag taaaattact
     3361 caagtacttt acacctctgg ttcttgaccc cctaccttca gcaagcccag cagatccact
     3421 agttctagag cggccgccac cgcggtggag ctccagcttt tgttcccttt agtgagggtt
     3481 aattgcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     3541 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     3601 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     3661 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     3721 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     3781 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     3841 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     3901 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     3961 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     4021 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     4081 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     4141 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     4201 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     4261 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     4321 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     4381 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     4441 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     4501 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaaggcat
     4561 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     4621 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     4681 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     4741 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     4801 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     4861 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     4921 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     4981 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     5041 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     5101 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     5161 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     5221 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     5281 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     5341 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     5401 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     5461 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     5521 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     5581 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     5641 ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     5701 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     5761 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     5821 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacgc
     5881 tggctaagaa ctcatcagcc tccccggtcc atctacccac gtaccaatgc accaattggc
     5941 cacaatgacg gctactacat ggtgccattc cttcctcttt ataggaatgg agactacctc
     6001 ctgtccaaca atgctcttgg atacgagtac gcctacctgt tggacccagg tcattgcaca
     6061 acaccagaaa tgccctctga tctgcaaaag acgtgaatat ctgttcagac acccatatcc
     6121 actctgttcc acacaggtca gaggtttgtc caggagttct tgacagaggt gtaaaaagta
     6181 ctcaaaaatt ttactcaagt gaaagtacaa gtacttaggg aaaattttac tcaattaaaa
     6241 gtaaaagtat ctggctagaa tcttacttga gtaaaagtaa aaaagtactc cattaaaatt
     6301 gtacttgagt attaaggaag taaaagtaaa agcaagaaag aaaactagag attcttgttt
     6361 aagcttttaa tctcaaaaaa cattaaatga aatgcataca aggttttatc ctgctttaga
     6421 actgtttgta tttaattatc aaactataag acagacaatc taatgccagt acacgctact
     6481 caaagttgta aaacctcaga tttaacttca gtagaagctg attctcaaaa ttgttagtgt
     6541 caagcctagc tcttttgggg ctgaaaagca atcctgcagt gctgaaaagc ctctcacagg
     6601 cagccgatgc gggaagaggt gtattagtct tgatagagag gctgcaaata gcaggaaacg
     6661 tgagcagaga ctccctggtg tctgaaacac aggcccattt acgccttaag atacattgat
     6721 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
     6781 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     6841 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa
     6901 aacctctaca aatgtggtat ggctgattat gatctagagt cgcggccgct ttacttgtac
     6961 agctcgtcca tgccgccggt ggagtggtgg ccctcggcgc gctcgtactg ttccacgatg
     7021 gtgtagtcct cgttgtggga ggtgatgtcc agcttgatgt tgacgttgta ggcgccgggc
     7081 agctgcacgg gcttcttggc cttgtaggtg gtcttgacct cggcgtcgta gtggccgccg
     7141 tccttcagct tcagcctctg cttgatctcg cccttcaggg cgccgtcctc ggggtacatc
     7201 cgctcggagg aggcctccca gcccatggtc ttcttctgca ttacggggcc gtcggagggg
     7261 aagttggtgc cgcgcagctt caccttgtag atgaactcgc cgtcctgcag ggaggagtcc
     7321 tgggtcacgg tcaccacgcc gccgtcctcg aagttcatca cgcgctccca cttgaagccc
     7381 tcggggaagg acagcttcaa gtagtcgggg atgtcggcgg ggtgcttcac gtaggccttg
     7441 gagccgtaca tgaactgagg ggacaggatg tcccaggcga agggcagggg gccgcccttg
     7501 gtcaccttca gcttggcggt ctgggtgccc tcgtaggggc ggccctcgcc ctcgccctcg
     7561 atctcgaact cgtggccgtt cacggagccc tccatgtgca ccttgaagcg catgaactcc
     7621 ttgatgatgg ccatgttgtc atcctcgccc ttgctcacca tgg
//
