LOCUS       sequence-174760-        7862 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  contains clec14a enhancer sequence, and clec14a specific gene 
            promoter driving expression of eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7862)
  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 7862)
  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;clecl14aCE1-P1Egfp
FEATURES             Location/Qualifiers
     source          1..7862
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(144..165)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(514..525,526..688)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    1057..1178
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(1301..2011)
                     /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(1381..1404)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(1579..1598)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1848..1866
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      2008..2017
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     2659..2675
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2695..2715
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    3223..3247
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      3900..3909
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3906..3908,3909..3911,3912..4622)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3906..3908,3909..3911,3912..4622)
                     /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(3951..3972)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4212..4231)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4559..4580
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    4623..4688
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(4709..4733)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(4759..4779)
                     /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(4787..4803)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    4819..4953
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(5392..5579,5580..5591)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(5625..5641)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5625..5641)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(5678..5698)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5678..5696)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5717..5733)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5730..5752)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5741..5757
                     /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(5765..5771,5772..5789,5790..5795))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5810..5831
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5948..5965)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6119..6707)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6199..6218)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6878..7669,7670..7738))
                     /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     7501..7520
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7739..7843)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
       61 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
      121 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
      181 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacgc tggctaagaa
      241 ctcatcagcc tccccggtcc atctacccac gtaccaatgc accaattggc cacaatgacg
      301 gctactacat ggtgccattc cttcctcttt ataggaatgg agactacctc ctgtccaaca
      361 aggctcttgg atacgagtac gcctacctgt tggacccagg tcattgcaca acaccagaaa
      421 tgccctctga tctgcaaaag acgtgaatat ctgttcagac acccatatcc actctgttcc
      481 acacaggtca gaggtttgtc caggagttct tgacagaggt gtaaaaagta ctcaaaaatt
      541 ttactcaagt gaaagtacaa gtacttaggg aaaattttac tcaattaaaa gtaaaagtat
      601 ctggctagaa tcttacttga gtaaaagtaa aaaagtactc cattaaaatt gtacttgagt
      661 attaaggaag taaaagtaaa agcaagaaag aaaactagag attcttgttt aagcttttaa
      721 tctcaaaaaa cattaaatga aatgcataca aggttttatc ctgctttaga actgtttgta
      781 tttaattatc aaactataag acagacaatc taatgccagt acacgctact caaagttgta
      841 aaacctcaga tttaacttca gtagaagctg attctcaaaa ttgttagtgt caagcctagc
      901 tcttttgggg ctgaaaagca atcctgcagt gctgaaaagc ctctcacagg cagccgatgc
      961 gggaagaggt gtattagtct tgatagagag gctgcaaata gcaggaaacg tgagcagaga
     1021 ctccctggtg tctgaaacac aggcccattt acgccttaag atacattgat gagtttggac
     1081 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     1141 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     1201 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
     1261 aatgtggtat ggctgattat gatctagagt cgcggccgct ttacttgtac agctcgtcca
     1321 tgccgccggt ggagtggtgg ccctcggcgc gctcgtactg ttccacgatg gtgtagtcct
     1381 cgttgtggga ggtgatgtcc agcttgatgt tgacgttgta ggcgccgggc agctgcacgg
     1441 gcttcttggc cttgtaggtg gtcttgacct cggcgtcgta gtggccgccg tccttcagct
     1501 tcagcctctg cttgatctcg cccttcaggg cgccgtcctc ggggtacatc cgctcggagg
     1561 aggcctccca gcccatggtc ttcttctgca ttacggggcc gtcggagggg aagttggtgc
     1621 cgcgcagctt caccttgtag atgaactcgc cgtcctgcag ggaggagtcc tgggtcacgg
     1681 tcaccacgcc gccgtcctcg aagttcatca cgcgctccca cttgaagccc tcggggaagg
     1741 acagcttcaa gtagtcgggg atgtcggcgg ggtgcttcac gtaggccttg gagccgtaca
     1801 tgaactgagg ggacaggatg tcccaggcga agggcagggg gccgcccttg gtcaccttca
     1861 gcttggcggt ctgggtgccc tcgtaggggc ggccctcgcc ctcgccctcg atctcgaact
     1921 cgtggccgtt cacggagccc tccatgtgca ccttgaagcg catgaactcc ttgatgatgg
     1981 ccatgttgtc atcctcgccc ttgctcacca tggtggcgac cggtaatgtc agacctggta
     2041 actccttact gtaacttatt gcacctgatg aacaaaagat gtgaactgaa cgcaagtagg
     2101 aggctaatag aggctagtac agtacaaaga tccccaaatg atgtgtgccg gccttagtgg
     2161 aggcagaccc ctggttatat acctgtgctg cagaaagagg ggcttacaaa tggatctctc
     2221 tggaatggca ttatctcatg gtgggaacag tgttgtcagc agaatcagcg gggtggactc
     2281 tgacctgaga atgacagtct agcactgaat ggctcagaca atactcttgt ggccaactat
     2341 caccggaaaa taagtgcctg ccaaaacaaa ccagaccaga tgaagctaca tggctgtaga
     2401 acgtcagtat ttaatgtgtt gtggccactt ggttcaccag atttaagagc taattttttt
     2461 aaatatcaca gataatcaat cgcttttttt tttcttgact gaaaagctta gactggaaaa
     2521 ttctttgaaa aagtttgcac tacagaaaat aaatacataa catcctgcac tgaatgcaca
     2581 ctattaatag aaaagagagc attattgtag gaaatgtatt gtatatgtac attctgtgca
     2641 tttcacaatt ttctcgagca ggaaacagct atgaccatga ttacgccaag ctatcaactt
     2701 tgtatagaaa agttgtcatt gttgtttaaa aaagcagcga gctctcgtaa taaaacacct
     2761 gtatcacctt catttcaact ttccttcttt gttctttcta ggcatacttg tagctccttt
     2821 tcctgttacc cataattcca tttcctgtcc ttcttcccca cccccctggc tctctagact
     2881 gcagaacaga gagtcgagtg ctcctgcaac ctcacttcct gccattcaga gccgtttcct
     2941 ccaggaagca tctgctgctc tctcatgaag cggcctgttg gaggaacaga ataataatac
     3001 ggtgttagtc atgggacatt caacaccaca ttagtcccat gtgagctctt gaaaacacac
     3061 gcggaggtca atcaacacgc atttgaaact gacagcgttg agtgcgacat gtaaacacag
     3121 aggtatagta ctactgtaaa gaacaatccc tgatgtaatg agcaacgctt cctggtattt
     3181 ttcccagagt tgttttgtta cagacacaat gaacaagtca aagcaagttt gtacaaaaaa
     3241 gcaggctcgc tagttgactt gacatatatg cacgtaatgt gttctgtaaa tatacaataa
     3301 tttattttcc acttaaagac ccttaaaaat agggcactga tgtaaagtac atttttaaat
     3361 ggcgaggtat aaagtggaat aattttcaaa actttatact cactactctt gaatattttt
     3421 agaagtgcta ctttttactc atattttgag tagcaggtac ttttacttgc actacatatt
     3481 tttgggaagt aaagatactt ttacttgagc atgatttttc agtcctcttt ccaccactgg
     3541 ctacgatgaa acctgaaacc ttcaaggaga tgatgggtgt ggatgcatga acggtttatc
     3601 agccctatgc caagaaacac aggtttcctc gtcatttaaa gctcatccgt cactctgctg
     3661 tgggagtttg gcagtgggat tgtgtattat ggaagcttca gtttcaccag gaacagattt
     3721 ttgttcatag ttgcgattgt tttgcttcgg cagcaccacc acccaagagg aagcgaatgg
     3781 ctgtatcaag gctgaacaga cttcctgtgc ttgttgtctt gggaggacct cacagggacg
     3841 gctaagaaag ccagggggag aaaaagcaga caatatcatt ttagtttgag tctgcgatcg
     3901 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     3961 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     4021 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     4081 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     4141 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     4201 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     4261 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     4321 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     4381 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     4441 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     4501 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     4561 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     4621 agtccggact cagatctcga gctcaagctt cgaattctgc agtcgacggt accgcgggcc
     4681 cgggatccac cggatctaga taactgatac ccagctttct tgtacaaagt ggggcgcgcc
     4741 tacgtaacta gtctcgagca actttattat acatagttga taattcactg gccgtcgttt
     4801 tacggtacca tcgatgatga tccagacatg ataagataca ttgatgagtt tggacaaacc
     4861 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     4921 tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg
     4981 tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt
     5041 ggtatggctg attatgatcc tctagatcag atctgcgaag atacggccac gggtgctctt
     5101 gatcctgtgg ctgattttgg actgtgctgc tcgcagctgc tgatgaatca catacttcct
     5161 ccattttctt ccactgattg actgttataa tttccctaat ttccaggtca aggtgctgtg
     5221 cattgtggta atagatgtga catgacgtca cttccaaagg accaatgaac atgtctgacc
     5281 aatttcatat aatgtgaaaa cgattttcat aggcagaata aataacattt aaattaaact
     5341 gggcatcagc gcaattcaat tggtttggta atagcaaggg aaaatagaat gaagtgatct
     5401 ccaaaaaata agtacttttt gactgtaaat aaaattgtaa ggagtaaaaa gtactttttt
     5461 ttctaaaaaa atgtaattaa gtaaaagtaa aagtattgat ttttaattgt actcaagtaa
     5521 agtaaaaatc cccaaaaata atacttaagt acagtaatca agtaaaatta ctcaagtact
     5581 ttacacctct ggttcttgac cccctacctt cagcaagccc agcagatcca ctagttctag
     5641 agcggccgcc accgcggtgg agctccagct tttgttccct ttagtgaggg ttaattgcgc
     5701 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     5761 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     5821 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     5881 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     5941 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     6001 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     6061 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     6121 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     6181 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     6241 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     6301 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     6361 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     6421 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     6481 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     6541 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     6601 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     6661 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     6721 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     6781 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     6841 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     6901 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     6961 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     7021 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     7081 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     7141 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     7201 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     7261 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     7321 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     7381 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     7441 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     7501 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     7561 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     7621 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     7681 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     7741 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     7801 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     7861 tg
//
