LOCUS       sequence-178859-        7834 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  contains lmo2 enhancer sequence, and lmo2 specific gene promoter 
            driving expression of eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7834)
  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 7834)
  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;lmo2CE1-P1Egfp
FEATURES             Location/Qualifiers
     source          1..7834
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     625..641
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    661..681
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    1148..1172
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      1838..1847
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1844..1846,1847..1849,1850..2560)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1844..1846,1847..1849,1850..2560)
                     /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(1889..1910)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2150..2169)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2497..2518
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2561..2626
                     /label=MCS
                     /note="multiple cloning site"
     protein_bind    complement(2647..2671)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2697..2717)
                     /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(2725..2741)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     polyA_signal    2757..2891
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    join(3330..3517,3518..3529)
                     /label=Tol2 left end
                     /note="left end of the Tol2 transposon from the medaka fish
                     Oryzias latipes (Kawakami, 2007)"
     primer_bind     complement(3563..3579)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3563..3579)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(3616..3636)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3616..3634)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3655..3671)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3668..3690)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3679..3695
                     /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(3703..3709,3710..3727,3728..3733))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3748..3769
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3886..3903)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4057..4645)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4137..4156)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4816..5607,5608..5676))
                     /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     5439..5458
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5677..5781)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5944..5965)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     misc_feature    join(6314..6325,6326..6488)
                     /label=Tol2 right end
                     /note="right end of the Tol2 transposon from the medaka 
                     fish Oryzias latipes (Kawakami, 2007)"
     polyA_signal    6857..6978
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(7101..7811)
                     /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(7181..7204)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(7379..7398)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     7648..7666
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      7808..7817
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 ctggtaactc cttactgtaa cttattgcac ctgatgaaca aaagatgtga actgaacgca
       61 agtaggaggc taatagaggc tagtacagta caaagatccc caaatgatgt gtgccggcct
      121 tagtggaggc agacccctgg ttatatacct gtgctgcaga aagaggggct tacaaatgga
      181 tctctctgga atggcattat ctcatggtgg gaacagtgtt gtcagcagaa tcagcggggt
      241 ggactctgac ctgagaatga cagtctagca ctgaatggct cagacaatac tcttgtggcc
      301 aactatcacc ggaaaataag tgcctgccaa aacaaaccag accagatgaa gctacatggc
      361 tgtagaacgt cagtatttaa tgtgttgtgg ccacttggtt caccagattt aagagctaat
      421 ttttttaaat atcacagata atcaatcgct tttttttttc ttgactgaaa agcttagact
      481 ggaaaattct ttgaaaaagt ttgcactaca gaaaataaat acataacatc ctgcactgaa
      541 tgcacactat taatagaaaa gagagcatta ttgtaggaaa tgtattgtat atgtacattc
      601 tgtgcatttc acaattttct cgagcaggaa acagctatga ccatgattac gccaagctat
      661 caactttgta tagaaaagtt gcaaaaagtc atcatggcca acagaatgtt atgtggaaga
      721 atgtgatcta atgtcaaaga aaatgattct cgtgacacat cgggagtatt gcatgacaat
      781 gaccagaaaa aaaaacgcta ataacagaga tgtcataagc catgttgctg tacagcattt
      841 accacacccc taaccctttt tgaagattta tgagttaatg gttttcctgc agtagatgag
      901 ttgacggttt ccaaggctct gtttcagaag ccatccttct attaaaggag ccttcagcca
      961 atcacagtcc agcagggcaa tgaagtgaag cgaacaggaa gtcaacacga ctcttacacc
     1021 ccctgccagg gttcatctgt gctcatttcc tgcacatcct ctggctttgc tgtgtccaat
     1081 ccacaggaaa cggtggacgc cttatgcaca cagtttaaca tttatttaaa acaaactgta
     1141 gaagcaaaca agtttgtaca aaaaagcagg ctcgctactg ctccagcaag agttctgaat
     1201 acataaaaaa gtaaaatgat tgtttaaccc taaaagcaat tataaaacaa taggcctatt
     1261 tgccaacaag acaaaatgtg gagagttctt tttaaggggt tcatccaaaa aaaaatctga
     1321 ttatgtcatt tacttttatt gttgcaaact gttgaacaca aacaaaaagt tgcaatgaat
     1381 cctgaaaacc agtattggct tccaaaatag cctattgtta cagttttcta acattcctca
     1441 caataacttt gtcaagagaa caaaacgaaa tgtttgccag ccaagttgtt taacctcctc
     1501 gtttggatgc agccaggtat ttgatttctg cctgactgac tccgtgtttg agctcgtgac
     1561 cccttaagtc acgagtcgtc ggtgtcggct gtataattga ctggtgattc gcgcatgtag
     1621 gagcgctatt tcctctcatt gtcgagccgt cgtgagcagg aagtcggaaa agagcagaaa
     1681 ctgagggggc agggccattt aaaaatacct gctgccctac aataacggga cacagagaca
     1741 gcagccctgc gggacactgg cgtggcgact tgcagcgcaa aatgcaccac aaacaagacg
     1801 gagcctaaac gccacattta tcacgcgccg gagaatcgcc accatggtga gcaagggcga
     1861 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     1921 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     1981 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
     2041 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
     2101 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     2161 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     2221 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     2281 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
     2341 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
     2401 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
     2461 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     2521 cgccgccggg atcactctcg gcatggacga gctgtacaag tccggactca gatctcgagc
     2581 tcaagcttcg aattctgcag tcgacggtac cgcgggcccg ggatccaccg gatctagata
     2641 actgataccc agctttcttg tacaaagtgg ggcgcgccta cgtaactagt ctcgagcaac
     2701 tttattatac atagttgata attcactggc cgtcgtttta cggtaccatc gatgatgatc
     2761 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     2821 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     2881 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
     2941 gggaggtttt ttaaagcaag taaaacctct acaaatgtgg tatggctgat tatgatcctc
     3001 tagatcagat ctgcgaagat acggccacgg gtgctcttga tcctgtggct gattttggac
     3061 tgtgctgctc gcagctgctg atgaatcaca tacttcctcc attttcttcc actgattgac
     3121 tgttataatt tccctaattt ccaggtcaag gtgctgtgca ttgtggtaat agatgtgaca
     3181 tgacgtcact tccaaaggac caatgaacat gtctgaccaa tttcatataa tgtgaaaacg
     3241 attttcatag gcagaataaa taacatttaa attaaactgg gcatcagcgc aattcaattg
     3301 gtttggtaat agcaagggaa aatagaatga agtgatctcc aaaaaataag tactttttga
     3361 ctgtaaataa aattgtaagg agtaaaaagt actttttttt ctaaaaaaat gtaattaagt
     3421 aaaagtaaaa gtattgattt ttaattgtac tcaagtaaag taaaaatccc caaaaataat
     3481 acttaagtac agtaatcaag taaaattact caagtacttt acacctctgg ttcttgaccc
     3541 cctaccttca gcaagcccag cagatccact agttctagag cggccgccac cgcggtggag
     3601 ctccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat catggtcata
     3661 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     3721 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     3781 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     3841 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     3901 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     3961 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     4021 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     4081 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     4141 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     4201 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     4261 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     4321 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4381 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4441 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     4501 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     4561 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     4621 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     4681 tcagtggaac gaaaactcac gttaaggcat tttggtcatg agattatcaa aaaggatctt
     4741 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     4801 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     4861 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     4921 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     4981 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     5041 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     5101 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     5161 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     5221 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     5281 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     5341 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     5401 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     5461 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     5521 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     5581 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     5641 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     5701 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     5761 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt
     5821 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     5881 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     5941 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     6001 aaaaccgtct atcagggcga tggcccacgc tggctaagaa ctcatcagcc tccccggtcc
     6061 atctacccac gtaccaatgc accaattggc cacaatgacg gctactacat ggtgccattc
     6121 cttcctcttt ataggaatgg agactacctc ctgtccaaca atgctcttgg atacgagtac
     6181 gcctacctgt tggacccagg tcattgcaca acaccagaaa tgccctctga tctgcaaaag
     6241 acgtgaatat ctgttcagac acccatatcc actctgttcc acacaggtca gaggtttgtc
     6301 caggagttct tgacagaggt gtaaaaagta ctcaaaaatt ttactcaagt gaaagtacaa
     6361 gtacttaggg aaaattttac tcaattaaaa gtaaaagtat ctggctagaa tcttacttga
     6421 gtaaaagtaa aaaagtactc cattaaaatt gtacttgagt attaaggaag taaaagtaaa
     6481 agcaagaaag aaaactagag attcttgttt aagcttttaa tctcaaaaaa cattaaatga
     6541 aatgcataca aggttttatc ctgctttaga actgtttgta tttaattatc aaactataag
     6601 acagacaatc taatgccagt acacgctact caaagttgta aaacctcaga tttaacttca
     6661 gtagaagctg attctcaaaa ttgttagtgt caagcctagc tcttttgggg ctgaaaagca
     6721 atcctgcagt gctgaaaagc ctctcacagg cagccgatgc gggaagaggt gtattagtct
     6781 tgatagagag gctgcaaata gcaggaaacg tgagcagaga ctccctggtg tctgaaacac
     6841 aggcccattt acgccttaag atacattgat gagtttggac aaaccacaac tagaatgcag
     6901 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     6961 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     7021 gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat ggctgattat
     7081 gatctagagt cgcggccgct ttacttgtac agctcgtcca tgccgccggt ggagtggtgg
     7141 ccctcggcgc gctcgtactg ttccacgatg gtgtagtcct cgttgtggga ggtgatgtcc
     7201 agcttgatgt tgacgttgta ggcgccgggc agctgcacgg gcttcttggc cttgtaggtg
     7261 gtcttgacct cggcgtcgta gtggccgccg tccttcagct tcagcctctg cttgatctcg
     7321 cccttcaggg cgccgtcctc ggggtacatc cgctcggagg aggcctccca gcccatggtc
     7381 ttcttctgca ttacggggcc gtcggagggg aagttggtgc cgcgcagctt caccttgtag
     7441 atgaactcgc cgtcctgcag ggaggagtcc tgggtcacgg tcaccacgcc gccgtcctcg
     7501 aagttcatca cgcgctccca cttgaagccc tcggggaagg acagcttcaa gtagtcgggg
     7561 atgtcggcgg ggtgcttcac gtaggccttg gagccgtaca tgaactgagg ggacaggatg
     7621 tcccaggcga agggcagggg gccgcccttg gtcaccttca gcttggcggt ctgggtgccc
     7681 tcgtaggggc ggccctcgcc ctcgccctcg atctcgaact cgtggccgtt cacggagccc
     7741 tccatgtgca ccttgaagcg catgaactcc ttgatgatgg ccatgttgtc atcctcgccc
     7801 ttgctcacca tggtggcgac cggtaatgtc agac
//
