LOCUS       sequence-178702-        7529 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  contains tmem88a enhancer 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 7529)
  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 7529)
  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;tmem88aCE1-basEgfp
FEATURES             Location/Qualifiers
     source          1..7529
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     644..660
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    680..700
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    1220..1244
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1321..1330
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1327..1329,1330..1332,1333..2043)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1327..1329,1330..1332,1333..2043)
                     /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(1372..1393)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1633..1652)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1980..2001
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2044..2109
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2233..2354
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(2361..2385)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2411..2431)
                     /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(2439..2455)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    2471..2605
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(3044..3231,3232..3243)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(3277..3293)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3277..3293)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3330..3350)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3330..3348)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3369..3385)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3382..3404)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3393..3409
                     /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(3417..3423,3424..3441,3442..3447))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3462..3483
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3600..3617)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3771..4359)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3851..3870)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4530..5321,5322..5390))
                     /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     5153..5172
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5391..5495)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5658..5679)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(6028..6039,6040..6202)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    6571..6692
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(6815..7525)
                     /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(6895..6918)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(7093..7112)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     7362..7380
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      join(7522..7529,1..2)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 gcgaccggta atgtcagacc tggtaactcc ttactgtaac ttattgcacc tgatgaacaa
       61 aagatgtgaa ctgaacgcaa gtaggaggct aatagaggct agtacagtac aaagatcccc
      121 aaatgatgtg tgccggcctt agtggaggca gacccctggt tatatacctg tgctgcagaa
      181 agaggggctt acaaatggat ctctctggaa tggcattatc tcatggtggg aacagtgttg
      241 tcagcagaat cagcggggtg gactctgacc tgagaatgac agtctagcac tgaatggctc
      301 agacaatact cttgtggcca actatcaccg gaaaataagt gcctgccaaa acaaaccaga
      361 ccagatgaag ctacatggct gtagaacgtc agtatttaat gtgttgtggc cacttggttc
      421 accagattta agagctaatt tttttaaata tcacagataa tcaatcgctt ttttttttct
      481 tgactgaaaa gcttagactg gaaaattctt tgaaaaagtt tgcactacag aaaataaata
      541 cataacatcc tgcactgaat gcacactatt aatagaaaag agagcattat tgtaggaaat
      601 gtattgtata tgtacattct gtgcatttca caattttctc gagcaggaaa cagctatgac
      661 catgattacg ccaagctatc aactttgtat agaaaagttg atttttgagt tattattcct
      721 gatataatga ttactataaa acaagactaa aagcttctga gtgtttacgt gtaagttatt
      781 tgtggattgt gaagtgtatt agaggaaggt tacagatggt cagtctcctg tctgtctgtg
      841 ggctggaagt gaggctcgta gaggtcatgg gacaggacta atgttttggt gacacctcgg
      901 ctctctctca ggactttact gcggaaatcc tggctccaag gccaggaagg caggttttcc
      961 tcttttatga ctctcaggat atatctcgtt tttcaccgtc cctctgcact taacgcttat
     1021 gttccaccag cagcccttta tctgtggctg ctgataaccc acagctgtga gtaaataaat
     1081 aaacaactgt caatagctta tttacttaca gataaagtcg ccatcctgtt tttcactctt
     1141 atgaactcga tatgttgttt tgttgactct atcacagtcg ttcacaactg gcaggccgtg
     1201 acctgaaaaa cattcaaagg caagtttgta caaaaaagca ggctagactc tagagggtat
     1261 ataatggatc tcgcgtctca gcctcacttt gagctcctcc acacgctagc gctaccggtc
     1321 gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
     1381 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc
     1441 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     1501 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
     1561 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
     1621 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
     1681 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     1741 gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag
     1801 aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
     1861 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     1921 aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
     1981 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
     2041 aagtccggac tcagatctcg agctcaagct tcgaattctg cagtcgacgg taccgcgggc
     2101 ccgggatcca ccggatctag ataactgatc ataatcagcc ataccacatt tgtagaggtt
     2161 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     2221 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     2281 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     2341 atcaatgtat cttaacgcgt acccagcttt cttgtacaaa gtggggcgcg cctacgtaac
     2401 tagtctcgag caactttatt atacatagtt gataattcac tggccgtcgt tttacggtac
     2461 catcgatgat gatccagaca tgataagata cattgatgag tttggacaaa ccacaactag
     2521 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac
     2581 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt
     2641 tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtatggc
     2701 tgattatgat cctctagatc agatctgcga agatacggcc acgggtgctc ttgatcctgt
     2761 ggctgatttt ggactgtgct gctcgcagct gctgatgaat cacatacttc ctccattttc
     2821 ttccactgat tgactgttat aatttcccta atttccaggt caaggtgctg tgcattgtgg
     2881 taatagatgt gacatgacgt cacttccaaa ggaccaatga acatgtctga ccaatttcat
     2941 ataatgtgaa aacgattttc ataggcagaa taaataacat ttaaattaaa ctgggcatca
     3001 gcgcaattca attggtttgg taatagcaag ggaaaataga atgaagtgat ctccaaaaaa
     3061 taagtacttt ttgactgtaa ataaaattgt aaggagtaaa aagtactttt ttttctaaaa
     3121 aaatgtaatt aagtaaaagt aaaagtattg atttttaatt gtactcaagt aaagtaaaaa
     3181 tccccaaaaa taatacttaa gtacagtaat caagtaaaat tactcaagta ctttacacct
     3241 ctggttcttg accccctacc ttcagcaagc ccagcagatc cactagttct agagcggccg
     3301 ccaccgcggt ggagctccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg
     3361 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     3421 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     3481 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     3541 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     3601 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3661 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3721 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3781 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3841 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3901 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3961 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4021 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4081 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4141 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4201 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4261 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4321 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4381 acggggtctg acgctcagtg gaacgaaaac tcacgttaag gcattttggt catgagatta
     4441 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4501 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4561 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4621 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4681 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4741 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4801 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4861 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4921 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4981 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     5041 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     5101 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     5161 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5221 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5281 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5341 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5401 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5461 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5521 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     5581 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     5641 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     5701 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc acgctggcta agaactcatc
     5761 agcctccccg gtccatctac ccacgtacca atgcaccaat tggccacaat gacggctact
     5821 acatggtgcc attccttcct ctttatagga atggagacta cctcctgtcc aacaatgctc
     5881 ttggatacga gtacgcctac ctgttggacc caggtcattg cacaacacca gaaatgccct
     5941 ctgatctgca aaagacgtga atatctgttc agacacccat atccactctg ttccacacag
     6001 gtcagaggtt tgtccaggag ttcttgacag aggtgtaaaa agtactcaaa aattttactc
     6061 aagtgaaagt acaagtactt agggaaaatt ttactcaatt aaaagtaaaa gtatctggct
     6121 agaatcttac ttgagtaaaa gtaaaaaagt actccattaa aattgtactt gagtattaag
     6181 gaagtaaaag taaaagcaag aaagaaaact agagattctt gtttaagctt ttaatctcaa
     6241 aaaacattaa atgaaatgca tacaaggttt tatcctgctt tagaactgtt tgtatttaat
     6301 tatcaaacta taagacagac aatctaatgc cagtacacgc tactcaaagt tgtaaaacct
     6361 cagatttaac ttcagtagaa gctgattctc aaaattgtta gtgtcaagcc tagctctttt
     6421 ggggctgaaa agcaatcctg cagtgctgaa aagcctctca caggcagccg atgcgggaag
     6481 aggtgtatta gtcttgatag agaggctgca aatagcagga aacgtgagca gagactccct
     6541 ggtgtctgaa acacaggccc atttacgcct taagatacat tgatgagttt ggacaaacca
     6601 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
     6661 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt
     6721 ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg
     6781 gtatggctga ttatgatcta gagtcgcggc cgctttactt gtacagctcg tccatgccgc
     6841 cggtggagtg gtggccctcg gcgcgctcgt actgttccac gatggtgtag tcctcgttgt
     6901 gggaggtgat gtccagcttg atgttgacgt tgtaggcgcc gggcagctgc acgggcttct
     6961 tggccttgta ggtggtcttg acctcggcgt cgtagtggcc gccgtccttc agcttcagcc
     7021 tctgcttgat ctcgcccttc agggcgccgt cctcggggta catccgctcg gaggaggcct
     7081 cccagcccat ggtcttcttc tgcattacgg ggccgtcgga ggggaagttg gtgccgcgca
     7141 gcttcacctt gtagatgaac tcgccgtcct gcagggagga gtcctgggtc acggtcacca
     7201 cgccgccgtc ctcgaagttc atcacgcgct cccacttgaa gccctcgggg aaggacagct
     7261 tcaagtagtc ggggatgtcg gcggggtgct tcacgtaggc cttggagccg tacatgaact
     7321 gaggggacag gatgtcccag gcgaagggca gggggccgcc cttggtcacc ttcagcttgg
     7381 cggtctgggt gccctcgtag gggcggccct cgccctcgcc ctcgatctcg aactcgtggc
     7441 cgttcacgga gccctccatg tgcaccttga agcgcatgaa ctccttgatg atggccatgt
     7501 tgtcatcctc gcccttgctc accatggtg
//
