LOCUS       sequence-446988-        6759 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 6759)
  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 6759)
  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 mR3-17c +SV40 base prom-GFP-IRES-AP
FEATURES             Location/Qualifiers
     source          1..6759
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     9..26
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(join(220..933,934..936,937..939))
                     /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             complement(join(223..933,934..936,937..939))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(265..286)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     614..633
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     873..894
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      936..945
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(980..1176)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      994..1129
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        1419..1523
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1524..1592,1593..2384)
                     /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     complement(1742..1761)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2555..3143
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3044..3063
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3297..3314
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    3382..3516
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3406..3425)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3460..3479
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(3522..3717)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3536..3671
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     protein_bind    3776..3797
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3812..3817,3818..3835,3836..3842)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3850..3866
                     /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)."
     primer_bind     3855..3877
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3874..3890
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3874..3890
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4233..4252
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    4251..4306
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     4352..4371
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(4866..6365)
                     /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"
     misc_feature    join(6377..6759,1..170)
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    join(6379..6759,1..193)
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(6539..6558)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
ORIGIN
        1 cagaccttgc attcctttgg cgagagggga aagaccccta ggaatgctcg tcaagaagac
       61 agggccaggt ttccgggccc tcacattgcc aaaagacggc aatatggtgg aaaataacat
      121 atagacaaac gcacaccggc cttattccaa gcggcttcgg ccagtaacgt tagggggggg
      181 ggagggagag gggggggggg ggggggggaa ttggccgctt tacttgtaca gctcgtccat
      241 gccgagagtg atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc
      301 gttggggtct ttgctcaggg cggactgggt gctcaggtag tggttgtcgg gcagcagcac
      361 ggggccgtcg ccgatggggg tgttctgctg gtagtggtcg gcgagctgca cgctgccgtc
      421 ctcgatgttg tggcggatct tgaagttcac cttgatgccg ttcttctgct tgtcggccat
      481 gatatagacg ttgtggctgt tgtagttgta ctccagcttg tgccccagga tgttgccgtc
      541 ctccttgaag tcgatgccct tcagctcgat gcggttcacc agggtgtcgc cctcgaactt
      601 cacctcggcg cgggtcttgt agttgccgtc gtccttgaag aagatggtgc gctcctggac
      661 gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt cggggtagcg
      721 gctgaagcac tgcacgccgt aggtcagggt ggtcacgagg gtgggccagg gcacgggcag
      781 cttgccggtg gtgcagatga acttcagggt cagcttgccg taggtggcat cgccctcgcc
      841 ctcgccggac acgctgaact tgtggccgtt tacgtcgccg tccagctcga ccaggatggg
      901 caccaccccg gtgaacagct cctcgccctt gctcaccatg gtggcgaccg gtggatcccg
      961 ggcccgcggt accaagcttt ttgcaaaagc ctaggcctcc aaaaaagcct cctcactact
     1021 tctggaatag ctcagaggcc gaggcggcct cggcctctgc ataaataaaa aaaattagtc
     1081 agcgatgggg cggagaatgg gcggaactgg gcggagttag gggcgggatg ggcggagtta
     1141 ggggcgggac tatggttgct gactaattga gatgcagatc gcagatctcg aggtttcttt
     1201 ctcttgtccc tccttctgta atctgacctc atcaagccct gggaaggctg atccaccgga
     1261 gcaaatcctt catccagagg gccaaaatca gcttaatttg tagcaatttg aagcaggtct
     1321 ttgccaggat ttgaatatca aattatccca tagcctagac aggcctggga gcttccccta
     1381 cccccgtcga cccaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     1441 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     1501 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     1561 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     1621 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     1681 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     1741 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     1801 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     1861 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     1921 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     1981 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     2041 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     2101 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     2161 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     2221 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     2281 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     2341 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     2401 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     2461 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     2521 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     2581 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     2641 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     2701 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     2761 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     2821 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     2881 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     2941 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     3001 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     3061 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     3121 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     3181 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     3241 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
     3301 agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt
     3361 ggccgattca ttaatgcagc ggatccagac atgataagat acattgatga gtttggacaa
     3421 accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct
     3481 ttatttgtaa ccattataag ctgcaataaa caagttagct ttttgcaaaa gcctaggcct
     3541 ccaaaaaagc ctcctcacta cttctggaat agctcagagg ccgaggcggc ctcggcctct
     3601 gcataaataa aaaaaattag tcagccatgg ggcggagaat gggcggaact gggcggagtt
     3661 aggggcggga tgggcggagt taggggcggg actatggttg ctgactaatt gagatgcgga
     3721 tccgctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     3781 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     3841 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
     3901 ccaagcttgg gctgcaggtc gagggatctc cataagagaa gagggacagc tatgactggg
     3961 agtagtcagg agaggaggaa aaatctggct agtaaaacat gtaaggaaaa ttttagggat
     4021 gttaaagaaa aaaataacac aaaacaaaat ataaaaaaaa tctaacctca agtcaaggct
     4081 tttctatgga ataaggaatg gacagcaggg ggctgtttca tatactgatg acctctttat
     4141 agccaacctt tgttcatggc agccagcata tgggcatatg ttgccaaact ctaaaccaaa
     4201 tactcattct gatgttttaa atgatttgcc ctcccatatg tccttccgag tgagagacac
     4261 aaaaaattcc aacacactat tgcaatgaaa ataaatttcc tttattagcc agaagtcaga
     4321 tgctcaaggg gcttcatgat gtccccataa tttttggcag agggaaaaag atctcagtgg
     4381 tatttgtgag ccagggcatt ggccacacca gccaccacct tctgataggc agcctgcacc
     4441 tgaggagtgc ggccggccgc gatatcctcg accccactgg gctgtaggtg gtggctggat
     4501 ggcctgagcc ccctcctttc ctggggtgca ggcagccaca gggcacaacc tgcagtccct
     4561 ggggttgggg tagggagtcc gagagcagcc aaggggcagg tttggtgtca tgggctcagt
     4621 ccccagagcg cacggggagg ggaaggttcc atggcaggac ctctgtatag ggactccggg
     4681 gatgacgact cgggctggag gccaggcagg acgacaggag gtggggagca ggagaactcc
     4741 gggatgggga agcaggagcc ccagggacgg gacactcagg gagcagtggc cgtctccagc
     4801 agcagcaggg tcccggccag cagaggaagc aacgcgggga ccacggaccg ccccgggtgc
     4861 gcggcgtcgg tggtgccggc ggggggcgcc aggtcgcagg cggtgtaggg ctccaggcag
     4921 gcggcgaagg ccatgacgtg cgctatgaag gtctgctcct gcacgccgtg aaccaggtgc
     4981 gcctgcgggc cgcgcgcgaa caccgccacg tcctcgcctg cgtgggtctc ttcgtccagg
     5041 ggcactgctg actgctgccg atactcgggg ctcccgctct cgctctcggt aacatccggc
     5101 cgggcgccgt ccttgagcac atagcctgga ccgtttccgt ataggaggac cgtgtaggcc
     5161 ttcctgtccc gggccttgcc aggggccagc ccgaagatgg agctccctcg cagggggtag
     5221 cctccgaagg agaagacgtg ggagtggtcg gcagtgacga ggctcagcgt gtcctcctcg
     5281 ctggtgagct ggcccgccct ctcaatggcg tcgtcgaaca tgatcgtctc agtcagtgcc
     5341 cggtaagccc tgctttcatg atgaccatgg tcgatgcgac caccctccac gaagaggaag
     5401 aagccgcggg ggttcctgct cagcaggcgc agggcagcct ctgtcatctc catcagggag
     5461 gggtccagtg tggagtctcg gtggatctcg tatttcatgt ctccaggctc aaagagaccc
     5521 atgagatggg tcacagacgg gtccagggaa gcctgcatga gctcagtgcg gttccacaca
     5581 taccgggcac cctggcgctt cgccagccat tcctgcacca gattcttccc gtccagcctg
     5641 gtcccacctt ggctgtagtc atctgggtac tcagggtctg gggttcccat gcgaaacatg
     5701 tactttcggc ctccacctag gatcacgtca atgtccatgt tggagatgag ctgcgtagcg
     5761 atgtcctggc acccctcctg gcgggccgag gcaggcacgt cggcgtccga gtaccagttg
     5821 cggttcaccg tgtgggcgta ggtgccggct ggcgaggcgt gctgcactcg tgtggtggtt
     5881 accactccca ctgacttccc tgctttcttg gcccgattca tcacggagat gacctcgttg
     5941 ccgcgtgtcg tgttgcactg gttaaagcgg gcggctgcac tcaagccaat ggtctggaag
     6001 ttgcccttga ccccgcacag gtaggccgtg gctgtggctc cactgtctgg cacatgtttg
     6061 tctacattgt atgtcttgga cagagccaca tatgggaagc ggtccatggc caggggtatc
     6121 tcaggcccca gtttgtcctt cttctgccct tttaggatcc tggcagctgt caccgtagac
     6181 acccccatcc catcgcccag gaagatgatg aggttcttgg cggctgtctg tgcaggctgc
     6241 agcttcttgg cggcacccag ggcctcggct gcctcgcggt tccagaagtc cgggttctcc
     6301 tcctcaactg ggatgatgcc cagggagagc tgtagcctca ggcccagcag cagcagcagc
     6361 agcagcctcg accccatatt atcatcgtgt ttttcaaagg aaaaccacgt ccccgtggtt
     6421 cggggggcct agacgttttt ttaacctcga ctaaacacat gtaaagcatg tgcaccgagg
     6481 ccccagatca gatcccatac aatggggtac cttctgggca tccttcagcc ccttgttgaa
     6541 tacgcttgag gagagccatt tgactctttc cacaactatc caactcacaa cgtggcactg
     6601 gggttgtgcc gcctttgcag gtgtatctta tacacgtggc ttttggccgc agaggcacct
     6661 gtcgccaggt ggggggttcc gctgcctgca aagggtcgct acagacgttg tttgtcttca
     6721 agaagcttcc agaggaactg cttccttcac gacattcaa
//
