LOCUS       sequence-178964-        7507 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  contains tmem88a specific gene promoter driving expression of eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7507)
  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 7507)
  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;tmem88aP1Egfp
FEATURES             Location/Qualifiers
     source          1..7507
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    11..35
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      674..683
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(680..682,683..685,686..1396)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(680..682,683..685,686..1396)
                     /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(725..746)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(986..1005)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1333..1354
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1397..1462
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(1483..1507)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1533..1553)
                     /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(1561..1577)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    1593..1727
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(2166..2353,2354..2365)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(2399..2415)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(2452..2470)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2491..2507)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2504..2526)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2515..2531
                     /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(2539..2545,2546..2563,2564..2569))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2584..2605
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2722..2739)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2893..3481)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2973..2992)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3652..4443,4444..4512))
                     /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     4275..4294
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4513..4617)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4780..4801)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(5150..5161,5162..5324)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    5693..5814
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(5937..6647)
                     /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(6017..6040)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(6215..6234)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     6484..6502
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      6644..6653
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     7295..7311
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7331..7351
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     primer_bind     7353..7372
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7353..7371
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    7380..7487
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     7396..7412
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     7397..7413
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7447..7463)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(join(7500..7507,1..11))
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 gagggttaat tcaagtttgt acaaaaaagc aggctcgcta aattaacaac ttctttccca
       61 acacctaaaa tatgcacatt ttcccatgaa atcttggcta aactacacat tttgtcagtt
      121 tgtgtgtgta tctgtaacac tttacagtaa agttgaacat tgtaaaaagc gattaggttc
      181 tgtacttcaa aattcagatt acttcatttt actttaagaa aagtgagtaa taatgtaagt
      241 atatctgaaa tgtattagca atttctgagc agaaataaaa catactgtgg agtgtgtaac
      301 tctaaagtca gtgcaagtca tagattttcc tactgcatgt gcaaaaaaat aaaaaagtct
      361 acaactaaac ccccagcaaa cccaacaccc cacatttctg aagggctgat ccagatcggg
      421 aggccaggag gagaagtggg agtggagaga tacgagtgag ttactgaggc aggacatctt
      481 cgtcaaagga agaaaagagt agagcaggaa agaaggaaga gaacgagctg acatagaaga
      541 ctgacgctca ttgagctcac agagcctgag gagatactac aggtggacct gaaaactgga
      601 taaaaatcaa gtcaaactaa ctgtagagta aagctatcat ctgccagtct gaggaatctg
      661 cccatatttc atcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      721 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
      781 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      841 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
      901 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      961 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     1021 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     1081 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
     1141 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
     1201 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     1261 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
     1321 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     1381 ggacgagctg tacaagtccg gactcagatc tcgagctcaa gcttcgaatt ctgcagtcga
     1441 cggtaccgcg ggcccgggat ccaccggatc tagataactg atacccagct ttcttgtaca
     1501 aagtggggcg cgcctacgta actagtctcg agcaacttta ttatacatag ttgataattc
     1561 actggccgtc gttttacggt accatcgatg atgatccaga catgataaga tacattgatg
     1621 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     1681 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     1741 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa
     1801 acctctacaa atgtggtatg gctgattatg atcctctaga tcagatctgc gaagatacgg
     1861 ccacgggtgc tcttgatcct gtggctgatt ttggactgtg ctgctcgcag ctgctgatga
     1921 atcacatact tcctccattt tcttccactg attgactgtt ataatttccc taatttccag
     1981 gtcaaggtgc tgtgcattgt ggtaatagat gtgacatgac gtcacttcca aaggaccaat
     2041 gaacatgtct gaccaatttc atataatgtg aaaacgattt tcataggcag aataaataac
     2101 atttaaatta aactgggcat cagcgcaatt caattggttt ggtaatagca agggaaaata
     2161 gaatgaagtg atctccaaaa aataagtact ttttgactgt aaataaaatt gtaaggagta
     2221 aaaagtactt ttttttctaa aaaaatgtaa ttaagtaaaa gtaaaagtat tgatttttaa
     2281 ttgtactcaa gtaaagtaaa aatccccaaa aataatactt aagtacagta atcaagtaaa
     2341 attactcaag tactttacac ctctggttct tgacccccta ccttcagcaa gcccagcaga
     2401 tccactagtt ctagagcggc cgccaccgcg gtggagctcc agcttttgtt ccctttagtg
     2461 agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     2521 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     2581 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     2641 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     2701 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     2761 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     2821 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     2881 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     2941 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     3001 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     3061 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     3121 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     3181 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     3241 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     3301 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     3361 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3421 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     3481 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     3541 aggcattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     3601 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     3661 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     3721 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     3781 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     3841 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     3901 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     3961 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     4021 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     4081 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     4141 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     4201 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     4261 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     4321 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     4381 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     4441 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     4501 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     4561 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     4621 atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa attcgcgtta
     4681 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     4741 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     4801 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     4861 ccacgctggc taagaactca tcagcctccc cggtccatct acccacgtac caatgcacca
     4921 attggccaca atgacggcta ctacatggtg ccattccttc ctctttatag gaatggagac
     4981 tacctcctgt ccaacaatgc tcttggatac gagtacgcct acctgttgga cccaggtcat
     5041 tgcacaacac cagaaatgcc ctctgatctg caaaagacgt gaatatctgt tcagacaccc
     5101 atatccactc tgttccacac aggtcagagg tttgtccagg agttcttgac agaggtgtaa
     5161 aaagtactca aaaattttac tcaagtgaaa gtacaagtac ttagggaaaa ttttactcaa
     5221 ttaaaagtaa aagtatctgg ctagaatctt acttgagtaa aagtaaaaaa gtactccatt
     5281 aaaattgtac ttgagtatta aggaagtaaa agtaaaagca agaaagaaaa ctagagattc
     5341 ttgtttaagc ttttaatctc aaaaaacatt aaatgaaatg catacaaggt tttatcctgc
     5401 tttagaactg tttgtattta attatcaaac tataagacag acaatctaat gccagtacac
     5461 gctactcaaa gttgtaaaac ctcagattta acttcagtag aagctgattc tcaaaattgt
     5521 tagtgtcaag cctagctctt ttggggctga aaagcaatcc tgcagtgctg aaaagcctct
     5581 cacaggcagc cgatgcggga agaggtgtat tagtcttgat agagaggctg caaatagcag
     5641 gaaacgtgag cagagactcc ctggtgtctg aaacacaggc ccatttacgc cttaagatac
     5701 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     5761 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     5821 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc
     5881 aagtaaaacc tctacaaatg tggtatggct gattatgatc tagagtcgcg gccgctttac
     5941 ttgtacagct cgtccatgcc gccggtggag tggtggccct cggcgcgctc gtactgttcc
     6001 acgatggtgt agtcctcgtt gtgggaggtg atgtccagct tgatgttgac gttgtaggcg
     6061 ccgggcagct gcacgggctt cttggccttg taggtggtct tgacctcggc gtcgtagtgg
     6121 ccgccgtcct tcagcttcag cctctgcttg atctcgccct tcagggcgcc gtcctcgggg
     6181 tacatccgct cggaggaggc ctcccagccc atggtcttct tctgcattac ggggccgtcg
     6241 gaggggaagt tggtgccgcg cagcttcacc ttgtagatga actcgccgtc ctgcagggag
     6301 gagtcctggg tcacggtcac cacgccgccg tcctcgaagt tcatcacgcg ctcccacttg
     6361 aagccctcgg ggaaggacag cttcaagtag tcggggatgt cggcggggtg cttcacgtag
     6421 gccttggagc cgtacatgaa ctgaggggac aggatgtccc aggcgaaggg cagggggccg
     6481 cccttggtca ccttcagctt ggcggtctgg gtgccctcgt aggggcggcc ctcgccctcg
     6541 ccctcgatct cgaactcgtg gccgttcacg gagccctcca tgtgcacctt gaagcgcatg
     6601 aactccttga tgatggccat gttgtcatcc tcgcccttgc tcaccatggt ggcgaccggt
     6661 aatgtcagac ctggtaactc cttactgtaa cttattgcac ctgatgaaca aaagatgtga
     6721 actgaacgca agtaggaggc taatagaggc tagtacagta caaagatccc caaatgatgt
     6781 gtgccggcct tagtggaggc agacccctgg ttatatacct gtgctgcaga aagaggggct
     6841 tacaaatgga tctctctgga atggcattat ctcatggtgg gaacagtgtt gtcagcagaa
     6901 tcagcggggt ggactctgac ctgagaatga cagtctagca ctgaatggct cagacaatac
     6961 tcttgtggcc aactatcacc ggaaaataag tgcctgccaa aacaaaccag accagatgaa
     7021 gctacatggc tgtagaacgt cagtatttaa tgtgttgtgg ccacttggtt caccagattt
     7081 aagagctaat ttttttaaat atcacagata atcaatcgct tttttttttc ttgactgaaa
     7141 agcttagact ggaaaattct ttgaaaaagt ttgcactaca gaaaataaat acataacatc
     7201 ctgcactgaa tgcacactat taatagaaaa gagagcatta ttgtaggaaa tgtattgtat
     7261 atgtacattc tgtgcatttc acaattttct cgagcaggaa acagctatga ccatgattac
     7321 gccaagctat caactttgta tagaaaagtt ggtaatacga ctcactatag ggcgaattgg
     7381 gtaccgggcc ccccctcgag gtcgacggta tcgataagct tgatatcgaa ttcctgcagc
     7441 ccgggggatc cactagttct agagcggccg ccaccgcggt ggagctccag cttttgttcc
     7501 ctttagt
//
