LOCUS       sequence-178942-        7521 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  contains clec14a specific gene promoter driving expression of eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7521)
  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 7521)
  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;clecl14aP1Egfp
FEATURES             Location/Qualifiers
     source          1..7521
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      680..689
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(686..688,689..691,692..1402)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(686..688,689..691,692..1402)
                     /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(731..752)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(992..1011)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1339..1360
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1403..1468
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(1489..1513)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1539..1559)
                     /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(1567..1583)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    1599..1733
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(2172..2359,2360..2371)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(2405..2421)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(2458..2476)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2497..2513)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2510..2532)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2521..2537
                     /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(2545..2551,2552..2569,2570..2575))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2590..2611
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2728..2745)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2899..3487)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2979..2998)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3658..4449,4450..4518))
                     /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     4281..4300
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4519..4623)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4786..4807)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(5156..5167,5168..5330)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    5699..5820
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(5943..6653)
                     /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(6023..6046)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(6221..6240)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     6490..6508
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      6650..6659
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     7301..7317
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7337..7357
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     7359..7378
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7359..7377
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    7386..7493
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     7402..7418
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     7403..7419
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7453..7469)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(join(7506..7521,1..3))
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 attcaagttt gtacaaaaaa gcaggctcgc tagttgactt gacatatatg cacgtaatgt
       61 gttctgtaaa tatacaataa tttattttcc acttaaagac ccttaaaaat agggcactga
      121 tgtaaagtac atttttaaat ggcgaggtat aaagtggaat aattttcaaa actttatact
      181 cactactctt gaatattttt agaagtgcta ctttttactc atattttgag tagcaggtac
      241 ttttacttgc actacatatt tttgggaagt aaagatactt ttacttgagc atgatttttc
      301 agtcctcttt ccaccactgg ctacgatgaa acctgaaacc ttcaaggaga tgatgggtgt
      361 ggatgcatga acggtttatc agccctatgc caagaaacac aggtttcctc gtcatttaaa
      421 gctcatccgt cactctgctg tgggagtttg gcagtgggat tgtgtattat ggaagcttca
      481 gtttcaccag gaacagattt ttgttcatag ttgcgattgt tttgcttcgg cagcaccacc
      541 acccaagagg aagcgaatgg ctgtatcaag gctgaacaga cttcctgtgc ttgttgtctt
      601 gggaggacct cacagggacg gctaagaaag ccagggggag aaaaagcaga caatatcatt
      661 ttagtttgag tctgcgatcg ccaccatggt gagcaagggc gaggagctgt tcaccggggt
      721 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
      781 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
      841 caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt
      901 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
      961 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
     1021 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
     1081 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
     1141 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     1201 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     1261 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc
     1321 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     1381 cggcatggac gagctgtaca agtccggact cagatctcga gctcaagctt cgaattctgc
     1441 agtcgacggt accgcgggcc cgggatccac cggatctaga taactgatac ccagctttct
     1501 tgtacaaagt ggggcgcgcc tacgtaacta gtctcgagca actttattat acatagttga
     1561 taattcactg gccgtcgttt tacggtacca tcgatgatga tccagacatg ataagataca
     1621 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
     1681 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca
     1741 acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca
     1801 agtaaaacct ctacaaatgt ggtatggctg attatgatcc tctagatcag atctgcgaag
     1861 atacggccac gggtgctctt gatcctgtgg ctgattttgg actgtgctgc tcgcagctgc
     1921 tgatgaatca catacttcct ccattttctt ccactgattg actgttataa tttccctaat
     1981 ttccaggtca aggtgctgtg cattgtggta atagatgtga catgacgtca cttccaaagg
     2041 accaatgaac atgtctgacc aatttcatat aatgtgaaaa cgattttcat aggcagaata
     2101 aataacattt aaattaaact gggcatcagc gcaattcaat tggtttggta atagcaaggg
     2161 aaaatagaat gaagtgatct ccaaaaaata agtacttttt gactgtaaat aaaattgtaa
     2221 ggagtaaaaa gtactttttt ttctaaaaaa atgtaattaa gtaaaagtaa aagtattgat
     2281 ttttaattgt actcaagtaa agtaaaaatc cccaaaaata atacttaagt acagtaatca
     2341 agtaaaatta ctcaagtact ttacacctct ggttcttgac cccctacctt cagcaagccc
     2401 agcagatcca ctagttctag agcggccgcc accgcggtgg agctccagct tttgttccct
     2461 ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     2521 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     2581 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     2641 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     2701 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     2761 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     2821 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     2881 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     2941 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     3001 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     3061 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     3121 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3181 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     3241 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3301 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     3361 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3421 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3481 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3541 acgttaaggc attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     3601 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     3661 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     3721 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     3781 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     3841 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     3901 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     3961 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     4021 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     4081 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     4141 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     4201 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     4261 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     4321 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     4381 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     4441 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     4501 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     4561 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     4621 gcgcacattt ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc
     4681 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc
     4741 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag
     4801 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc
     4861 gatggcccac gctggctaag aactcatcag cctccccggt ccatctaccc acgtaccaat
     4921 gcaccaattg gccacaatga cggctactac atggtgccat tccttcctct ttataggaat
     4981 ggagactacc tcctgtccaa caatgctctt ggatacgagt acgcctacct gttggaccca
     5041 ggtcattgca caacaccaga aatgccctct gatctgcaaa agacgtgaat atctgttcag
     5101 acacccatat ccactctgtt ccacacaggt cagaggtttg tccaggagtt cttgacagag
     5161 gtgtaaaaag tactcaaaaa ttttactcaa gtgaaagtac aagtacttag ggaaaatttt
     5221 actcaattaa aagtaaaagt atctggctag aatcttactt gagtaaaagt aaaaaagtac
     5281 tccattaaaa ttgtacttga gtattaagga agtaaaagta aaagcaagaa agaaaactag
     5341 agattcttgt ttaagctttt aatctcaaaa aacattaaat gaaatgcata caaggtttta
     5401 tcctgcttta gaactgtttg tatttaatta tcaaactata agacagacaa tctaatgcca
     5461 gtacacgcta ctcaaagttg taaaacctca gatttaactt cagtagaagc tgattctcaa
     5521 aattgttagt gtcaagccta gctcttttgg ggctgaaaag caatcctgca gtgctgaaaa
     5581 gcctctcaca ggcagccgat gcgggaagag gtgtattagt cttgatagag aggctgcaaa
     5641 tagcaggaaa cgtgagcaga gactccctgg tgtctgaaac acaggcccat ttacgcctta
     5701 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt
     5761 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt
     5821 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt
     5881 taaagcaagt aaaacctcta caaatgtggt atggctgatt atgatctaga gtcgcggccg
     5941 ctttacttgt acagctcgtc catgccgccg gtggagtggt ggccctcggc gcgctcgtac
     6001 tgttccacga tggtgtagtc ctcgttgtgg gaggtgatgt ccagcttgat gttgacgttg
     6061 taggcgccgg gcagctgcac gggcttcttg gccttgtagg tggtcttgac ctcggcgtcg
     6121 tagtggccgc cgtccttcag cttcagcctc tgcttgatct cgcccttcag ggcgccgtcc
     6181 tcggggtaca tccgctcgga ggaggcctcc cagcccatgg tcttcttctg cattacgggg
     6241 ccgtcggagg ggaagttggt gccgcgcagc ttcaccttgt agatgaactc gccgtcctgc
     6301 agggaggagt cctgggtcac ggtcaccacg ccgccgtcct cgaagttcat cacgcgctcc
     6361 cacttgaagc cctcggggaa ggacagcttc aagtagtcgg ggatgtcggc ggggtgcttc
     6421 acgtaggcct tggagccgta catgaactga ggggacagga tgtcccaggc gaagggcagg
     6481 gggccgccct tggtcacctt cagcttggcg gtctgggtgc cctcgtaggg gcggccctcg
     6541 ccctcgccct cgatctcgaa ctcgtggccg ttcacggagc cctccatgtg caccttgaag
     6601 cgcatgaact ccttgatgat ggccatgttg tcatcctcgc ccttgctcac catggtggcg
     6661 accggtaatg tcagacctgg taactcctta ctgtaactta ttgcacctga tgaacaaaag
     6721 atgtgaactg aacgcaagta ggaggctaat agaggctagt acagtacaaa gatccccaaa
     6781 tgatgtgtgc cggccttagt ggaggcagac ccctggttat atacctgtgc tgcagaaaga
     6841 ggggcttaca aatggatctc tctggaatgg cattatctca tggtgggaac agtgttgtca
     6901 gcagaatcag cggggtggac tctgacctga gaatgacagt ctagcactga atggctcaga
     6961 caatactctt gtggccaact atcaccggaa aataagtgcc tgccaaaaca aaccagacca
     7021 gatgaagcta catggctgta gaacgtcagt atttaatgtg ttgtggccac ttggttcacc
     7081 agatttaaga gctaattttt ttaaatatca cagataatca atcgcttttt tttttcttga
     7141 ctgaaaagct tagactggaa aattctttga aaaagtttgc actacagaaa ataaatacat
     7201 aacatcctgc actgaatgca cactattaat agaaaagaga gcattattgt aggaaatgta
     7261 ttgtatatgt acattctgtg catttcacaa ttttctcgag caggaaacag ctatgaccat
     7321 gattacgcca agctatcaac tttgtataga aaagttggta atacgactca ctatagggcg
     7381 aattgggtac cgggcccccc ctcgaggtcg acggtatcga taagcttgat atcgaattcc
     7441 tgcagcccgg gggatccact agttctagag cggccgccac cgcggtggag ctccagcttt
     7501 tgttcccttt agtgagggtt a
//
