LOCUS       sequence-446954-        7227 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Enhancer reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7227)
  AUTHORS   Honnell V, Sweeney S, Norrie J, Parks M, Ramirez C, Jannu AJ, Xu B, 
            Teubner B, Lee AY, Bell C, Dyer MA
  TITLE     Evolutionary conservation of VSX2 super-enhancer modules in retinal 
            development.
  JOURNAL   Development. 2024 Jul 1;151(13):dev202435. doi: 10.1242/dev.202435. 
            Epub 2024 Jul 12.
  PUBMED    38994775
REFERENCE   2  (bases 1 to 7227)
  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"; journalName: 
            "Development."; date: "2024-07-1"; volume: "151(13):dev202435. doi";
            pages: " 10.1242/dev.202435. Epub 2024 Jul 12"
COMMENT     Sequence Label: Vsx2 mR3-17d +SV40 base prom-GFP-IRES-AP
FEATURES             Location/Qualifiers
     source          1..7227
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        336..532
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      383..518
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      567..576
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(573..575,576..578,579..1292)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(573..575,576..578,579..1289)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(618..639)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(879..898)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1226..1247
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1319..1892
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    1342..1894
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1486..1503)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1713..1732
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             1906..3405
                     /codon_start=1
                     /product="secreted alkaline phosphatase from human 
                     placenta"
                     /label=SEAP
                     /note="The natural enzyme has a C-terminal transmembrane 
                     sequence that is replaced by a GPI anchor."
                     /translation="LLLLLLLGLRLQLSLGIIPVEEENPDFWNREAAEALGAAKKLQPA
                     QTAAKNLIIFLGDGMGVSTVTAARILKGQKKDKLGPEIPLAMDRFPYVALSKTYNVDKH
                     VPDSGATATAYLCGVKGNFQTIGLSAAARFNQCNTTRGNEVISVMNRAKKAGKSVGVVT
                     TTRVQHASPAGTYAHTVNRNWYSDADVPASARQEGCQDIATQLISNMDIDVILGGGRKY
                     MFRMGTPDPEYPDDYSQGGTRLDGKNLVQEWLAKRQGARYVWNRTELMQASLDPSVTHL
                     MGLFEPGDMKYEIHRDSTLDPSLMEMTEAALRLLSRNPRGFFLFVEGGRIDHGHHESRA
                     YRALTETIMFDDAIERAGQLTSEEDTLSLVTADHSHVFSFGGYPLRGSSIFGLAPGKAR
                     DRKAYTVLLYGNGPGYVLKDGARPDVTESESGSPEYRQQSAVPLDEETHAGEDVAVFAR
                     GPQAHLVHGVQEQTFIAHVMAFAACLEPYTACDLAPPAGTTD"
     primer_bind     complement(3900..3919)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3965..4020
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4019..4038)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4381..4397)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4381..4397)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4394..4416)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4405..4421
                     /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(4429..4435,4436..4453,4454..4459))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4474..4495
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4554..4749
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4600..4735
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4755..4889
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4792..4811)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4846..4865
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4957..4974)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5128..5716)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5208..5227)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5887..6678,6679..6747))
                     /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     6510..6529
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6748..6852)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gggggggggg gctccccatc tgagagttta tcagaaccta atccggggag aagagcactg
       61 gctggggctg cttgcccgct aatcccagct gccattaaaa tattaaagat aaatctaatc
      121 gtctctttat ccaaaataag cgacttttgt gtggggagaa aacgtctaac cccttaggag
      181 gagaattagt tctaatgcat caaatggaat tcggtccagg ctgaggcgaa atagggttta
      241 gaggcaaatg agacggtaat agaactaaaa cccagaatga caaattaaaa accctcattt
      301 acacgaatta aaacggctcc tcgagatctg cgatctgcat ctcaattagt cagcaaccat
      361 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
      421 gccccatcgc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
      481 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagcttgg
      541 taccgcgggc ccgggatcca ccggtcgcca ccatggtgag caagggcgag gagctgttca
      601 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
      661 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
      721 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
      781 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
      841 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
      901 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
      961 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     1021 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     1081 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     1141 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     1201 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     1261 tcactctcgg catggacgag ctgtacaagt aaagcggcca attccgcccc cccccccccc
     1321 cctctccctc ccccccccct aacgttactg gccgaagccg cttggaataa ggccggtgtg
     1381 cgtttgtcta tatgttattt tccaccatat tgccgtcttt tggcaatgtg agggcccgga
     1441 aacctggccc tgtcttcttg acgagcattc ctaggggtct ttcccctctc gccaaaggaa
     1501 tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct ggaagcttct tgaagacaaa
     1561 caacgtctgt agcgaccctt tgcaggcagc ggaacccccc acctggcgac aggtgcctct
     1621 gcggccaaaa gccacgtgta taagatacac ctgcaaaggc ggcacaaccc cagtgccacg
     1681 ttgtgagttg gatagttgtg gaaagagtca aatggctctc ctcaagcgta ttcaacaagg
     1741 ggctgaagga tgcccagaag gtaccccatt gtatgggatc tgatctgggg cctcggtgca
     1801 catgctttac atgtgtttag tcgaggttaa aaaaacgtct aggccccccg aaccacgggg
     1861 acgtggtttt cctttgaaaa acacgatgat aatatggggt cgaggctgct gctgctgctg
     1921 ctgctgggcc tgaggctaca gctctccctg ggcatcatcc cagttgagga ggagaacccg
     1981 gacttctgga accgcgaggc agccgaggcc ctgggtgccg ccaagaagct gcagcctgca
     2041 cagacagccg ccaagaacct catcatcttc ctgggcgatg ggatgggggt gtctacggtg
     2101 acagctgcca ggatcctaaa agggcagaag aaggacaaac tggggcctga gatacccctg
     2161 gccatggacc gcttcccata tgtggctctg tccaagacat acaatgtaga caaacatgtg
     2221 ccagacagtg gagccacagc cacggcctac ctgtgcgggg tcaagggcaa cttccagacc
     2281 attggcttga gtgcagccgc ccgctttaac cagtgcaaca cgacacgcgg caacgaggtc
     2341 atctccgtga tgaatcgggc caagaaagca gggaagtcag tgggagtggt aaccaccaca
     2401 cgagtgcagc acgcctcgcc agccggcacc tacgcccaca cggtgaaccg caactggtac
     2461 tcggacgccg acgtgcctgc ctcggcccgc caggaggggt gccaggacat cgctacgcag
     2521 ctcatctcca acatggacat tgacgtgatc ctaggtggag gccgaaagta catgtttcgc
     2581 atgggaaccc cagaccctga gtacccagat gactacagcc aaggtgggac caggctggac
     2641 gggaagaatc tggtgcagga atggctggcg aagcgccagg gtgcccggta tgtgtggaac
     2701 cgcactgagc tcatgcaggc ttccctggac ccgtctgtga cccatctcat gggtctcttt
     2761 gagcctggag acatgaaata cgagatccac cgagactcca cactggaccc ctccctgatg
     2821 gagatgacag aggctgccct gcgcctgctg agcaggaacc cccgcggctt cttcctcttc
     2881 gtggagggtg gtcgcatcga ccatggtcat catgaaagca gggcttaccg ggcactgact
     2941 gagacgatca tgttcgacga cgccattgag agggcgggcc agctcaccag cgaggaggac
     3001 acgctgagcc tcgtcactgc cgaccactcc cacgtcttct ccttcggagg ctaccccctg
     3061 cgagggagct ccatcttcgg gctggcccct ggcaaggccc gggacaggaa ggcctacacg
     3121 gtcctcctat acggaaacgg tccaggctat gtgctcaagg acggcgcccg gccggatgtt
     3181 accgagagcg agagcgggag ccccgagtat cggcagcagt cagcagtgcc cctggacgaa
     3241 gagacccacg caggcgagga cgtggcggtg ttcgcgcgcg gcccgcaggc gcacctggtt
     3301 cacggcgtgc aggagcagac cttcatagcg cacgtcatgg ccttcgccgc ctgcctggag
     3361 ccctacaccg cctgcgacct ggcgcccccc gccggcacca ccgacgccgc gcacccgggg
     3421 cggtccgtgg tccccgcgtt gcttcctctg ctggccggga ccctgctgct gctggagacg
     3481 gccactgctc cctgagtgtc ccgtccctgg ggctcctgct tccccatccc ggagttctcc
     3541 tgctccccac ctcctgtcgt cctgcctggc ctccagcccg agtcgtcatc cccggagtcc
     3601 ctatacagag gtcctgccat ggaaccttcc cctccccgtg cgctctgggg actgagccca
     3661 tgacaccaaa cctgcccctt ggctgctctc ggactcccta ccccaacccc agggactgca
     3721 ggttgtgccc tgtggctgcc tgcaccccag gaaaggaggg ggctcaggcc atccagccac
     3781 cacctacagc ccagtggggt cgaggatatc gcggccggcc gcactcctca ggtgcaggct
     3841 gcctatcaga aggtggtggc tggtgtggcc aatgccctgg ctcacaaata ccactgagat
     3901 ctttttccct ctgccaaaaa ttatggggac atcatgaagc cccttgagca tctgacttct
     3961 ggctaataaa ggaaatttat tttcattgca atagtgtgtt ggaatttttt gtgtctctca
     4021 ctcggaagga catatgggag ggcaaatcat ttaaaacatc agaatgagta tttggtttag
     4081 agtttggcaa catatgccca tatgctggct gccatgaaca aaggttggct ataaagaggt
     4141 catcagtata tgaaacagcc ccctgctgtc cattccttat tccatagaaa agccttgact
     4201 tgaggttaga ttttttttat attttgtttt gtgttatttt tttctttaac atccctaaaa
     4261 ttttccttac atgttttact agccagattt ttcctcctct cctgactact cccagtcata
     4321 gctgtccctc ttctcttatg gagatccctc gacctgcagc ccaagcttgg cgtaatcatg
     4381 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4441 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4501 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagcgga tccgcatctc
     4561 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     4621 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     4681 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     4741 ttttgcaaaa agctaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     4801 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     4861 tcatcaatgt atcttatcat gtctggatcc gctgcattaa tgaatcggcc aacgcgcggg
     4921 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4981 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5041 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5101 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5161 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5221 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5281 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     5341 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5401 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5461 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5521 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5581 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5641 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5701 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5761 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5821 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5881 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5941 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6001 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6061 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6121 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6181 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6241 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6301 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6361 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6421 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6481 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6541 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6601 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6661 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6721 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6781 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6841 aggggttccg cgcacatttc cccgaaaagt gccacctggg tcgactgtgg gtactcaggg
     6901 tcacactcag ttcttcaggg aagagtcaat cccggccttc tcctcactcc cccccacccg
     6961 gcagaacctc atctccatct gcttgatcag ggcctgcact gtatattgca gcaattttaa
     7021 attgcaccat taaagatttt atgctggaga cgttaaaatt aaaaagatct atgtgcaaag
     7081 agtcacaacg aatttaatca acttgattta atagaaaaca gcaaatggaa tttatgatgc
     7141 tccaaaagac aagggaattt ttttcccaca gaaaaataat aaaaaagcac tttgtagaaa
     7201 acagaaagtg gagtacgggg ggggggg
//
