LOCUS       sequence-446953-        8191 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 8191)
  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 8191)
  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 Mini-enhancer +SV40 base prom-GFP-IRES-AP
FEATURES             Location/Qualifiers
     source          1..8191
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    763..1315
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    765..1338
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(925..944)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     1154..1171
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(join(1363..2076,2077..2079,2080..2082))
                     /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(1366..2076,2077..2079,2080..2082))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1408..1429)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1757..1776
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2016..2037
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2079..2088
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2123..2319)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2137..2272
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3996..4100
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4101..4169,4170..4961)
                     /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(4319..4338)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5132..5720
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5621..5640
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5874..5891
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    5959..6093
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5983..6002)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6037..6056
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(6099..6294)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6113..6248
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     protein_bind    6353..6374
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6389..6394,6395..6412,6413..6419)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6427..6443
                     /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     6432..6454
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6451..6467
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6451..6467
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6810..6829
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    6828..6883
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     6929..6948
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(join(7443..8191,1..751))
                     /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"
ORIGIN
        1 gcaccagatt cttcccgtcc agcctggtcc caccttggct gtagtcatct gggtactcag
       61 ggtctggggt tcccatgcga aacatgtact ttcggcctcc acctaggatc acgtcaatgt
      121 ccatgttgga gatgagctgc gtagcgatgt cctggcaccc ctcctggcgg gccgaggcag
      181 gcacgtcggc gtccgagtac cagttgcggt tcaccgtgtg ggcgtaggtg ccggctggcg
      241 aggcgtgctg cactcgtgtg gtggttacca ctcccactga cttccctgct ttcttggccc
      301 gattcatcac ggagatgacc tcgttgccgc gtgtcgtgtt gcactggtta aagcgggcgg
      361 ctgcactcaa gccaatggtc tggaagttgc ccttgacccc gcacaggtag gccgtggctg
      421 tggctccact gtctggcaca tgtttgtcta cattgtatgt cttggacaga gccacatatg
      481 ggaagcggtc catggccagg ggtatctcag gccccagttt gtccttcttc tgccctttta
      541 ggatcctggc agctgtcacc gtagacaccc ccatcccatc gcccaggaag atgatgaggt
      601 tcttggcggc tgtctgtgca ggctgcagct tcttggcggc acccagggcc tcggctgcct
      661 cgcggttcca gaagtccggg ttctcctcct caactgggat gatgcccagg gagagctgta
      721 gcctcaggcc cagcagcagc agcagcagca gcctcgaccc catattatca tcgtgttttt
      781 caaaggaaaa ccacgtcccc gtggttcggg gggcctagac gtttttttaa cctcgactaa
      841 acacatgtaa agcatgtgca ccgaggcccc agatcagatc ccatacaatg gggtaccttc
      901 tgggcatcct tcagcccctt gttgaatacg cttgaggaga gccatttgac tctttccaca
      961 actatccaac tcacaacgtg gcactggggt tgtgccgcct ttgcaggtgt atcttataca
     1021 cgtggctttt ggccgcagag gcacctgtcg ccaggtgggg ggttccgctg cctgcaaagg
     1081 gtcgctacag acgttgtttg tcttcaagaa gcttccagag gaactgcttc cttcacgaca
     1141 ttcaacagac cttgcattcc tttggcgaga ggggaaagac ccctaggaat gctcgtcaag
     1201 aagacagggc caggtttccg ggccctcaca ttgccaaaag acggcaatat ggtggaaaat
     1261 aacatataga caaacgcaca ccggccttat tccaagcggc ttcggccagt aacgttaggg
     1321 gggggggagg gagagggggg gggggggggg gaattggccg ctttacttgt acagctcgtc
     1381 catgccgaga gtgatcccgg cggcggtcac gaactccagc aggaccatgt gatcgcgctt
     1441 ctcgttgggg tctttgctca gggcggactg ggtgctcagg tagtggttgt cgggcagcag
     1501 cacggggccg tcgccgatgg gggtgttctg ctggtagtgg tcggcgagct gcacgctgcc
     1561 gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg ccgttcttct gcttgtcggc
     1621 catgatatag acgttgtggc tgttgtagtt gtactccagc ttgtgcccca ggatgttgcc
     1681 gtcctccttg aagtcgatgc ccttcagctc gatgcggttc accagggtgt cgccctcgaa
     1741 cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg tgcgctcctg
     1801 gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt ggtcggggta
     1861 gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc agggcacggg
     1921 cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg catcgccctc
     1981 gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct cgaccaggat
     2041 gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc atggtggcga ccggtggatc
     2101 ccgggcccgc ggtaccaagc tttttgcaaa agcctaggcc tccaaaaaag cctcctcact
     2161 acttctggaa tagctcagag gccgaggcgg cctcggcctc tgcataaata aaaaaaatta
     2221 gtcagcgatg gggcggagaa tgggcggaac tgggcggagt taggggcggg atgggcggag
     2281 ttaggggcgg gactatggtt gctgactaat tgagatgcag atcgcagatc tcgaggagcc
     2341 gttttaattc gtgtaaatga gggtttttaa tttgtcattc tgggttttag ttctattacc
     2401 gtctcatttg cctctaaacc ctatttcgcc tcagcctgga ccgaattcca tttgatgcat
     2461 tagaactaat tctcctccta aggggttaga cgttttctcc ccacacaaaa gtcgcttatt
     2521 ttggataaag agacgattag atttatcttt aatattttaa tggcagctgg gattagcggg
     2581 caagcagccc cagccagtgc tcttctcccc ggattaggtt ctgataaact ctcagatggg
     2641 gagccccccc cccccgtact ccactttctg ttttctacaa agtgcttttt tattattttt
     2701 ctgtgggaaa aaaattccct tgtcttttgg agcatcataa attccatttg ctgttttcta
     2761 ttaaatcaag ttgattaaat tcgttgtgac tctttgcaca tagatctttt taattttaac
     2821 gtctccagca taaaatcttt aatggtgcaa tttaaaattg ctgcaatata cagtgcaggc
     2881 cctgatcaag cagatggaga tgaggttctg ccgggtgggg gggagtgagg agaaggccgg
     2941 gattgactct tccctgaaga actgagtgtg accctgagta cccacagttt ctttctcttg
     3001 tccctccttc tgtaatctga cctcatcaag ccctgggaag gctgatccac cggagcaaat
     3061 ccttcatcca gagggccaaa atcagcttaa tttgtagcaa tttgaagcag gtctttgcca
     3121 ggatttgaat atcaaattat cccatagcct agacaggcct gggagcttcc cctacccccg
     3181 tgcacagcac tgatcctaac caagtttgcc agcccagctt ttctcatggc tcccacacaa
     3241 tggtctgctt tgtgcccacc tgccgataat gcttaattag gcgaggagtt gattaatttg
     3301 aggtaattga cagctaatta gaagcctaca ggctcattaa cttgacatct tgtctttgtg
     3361 aattggctgg gtacggtcag gtgatggtaa ccgttgctcc cctccccagc tcttccaacc
     3421 ccaaagtctg aggcatcacc taggggaggg ctgggcagat cgaccgtgtt ccccatccct
     3481 cctccctgca tctgtcccct ggttagagtt gtatgaagcg gaagacaatc tggggcttat
     3541 tagtgtccca gggagacatg taactgtgct gctgggaccc aaacagcaag agtcttagca
     3601 gcaaactcca gccgagaaag gactgaggga ttgtctcacc taattacagg gaatggctct
     3661 ctttgtattt gtgattcggt acggttgtca gatcgctttt gatcgctttt ctccatttgc
     3721 tggctcaggc catgctggaa ttgtgctaat tagaggagaa atggacagag tgatcctgtc
     3781 aaccctggtt actggtgggg gacaaagcag aatattattc taaggcaatg ttttgtaaga
     3841 gcccttctcc tgctctcatt aggaggaaga ggccccagga ggggaagttc cgaatgcttg
     3901 gcctggctaa tggagcattg gggtgttctt gtgctaatta acaggctctg ccagcactcc
     3961 aggtcgaccc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     4021 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     4081 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     4141 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     4201 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     4261 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     4321 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     4381 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     4441 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     4501 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     4561 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     4621 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     4681 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4741 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4801 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     4861 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     4921 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     4981 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     5041 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5101 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5161 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5221 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     5281 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5341 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5401 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5461 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5521 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5581 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5641 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5701 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5761 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5821 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     5881 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     5941 cgattcatta atgcagcgga tccagacatg ataagataca ttgatgagtt tggacaaacc
     6001 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     6061 tttgtaacca ttataagctg caataaacaa gttagctttt tgcaaaagcc taggcctcca
     6121 aaaaagcctc ctcactactt ctggaatagc tcagaggccg aggcggcctc ggcctctgca
     6181 taaataaaaa aaattagtca gccatggggc ggagaatggg cggaactggg cggagttagg
     6241 ggcgggatgg gcggagttag gggcgggact atggttgctg actaattgag atgcggatcc
     6301 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     6361 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     6421 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
     6481 agcttgggct gcaggtcgag ggatctccat aagagaagag ggacagctat gactgggagt
     6541 agtcaggaga ggaggaaaaa tctggctagt aaaacatgta aggaaaattt tagggatgtt
     6601 aaagaaaaaa ataacacaaa acaaaatata aaaaaaatct aacctcaagt caaggctttt
     6661 ctatggaata aggaatggac agcagggggc tgtttcatat actgatgacc tctttatagc
     6721 caacctttgt tcatggcagc cagcatatgg gcatatgttg ccaaactcta aaccaaatac
     6781 tcattctgat gttttaaatg atttgccctc ccatatgtcc ttccgagtga gagacacaaa
     6841 aaattccaac acactattgc aatgaaaata aatttccttt attagccaga agtcagatgc
     6901 tcaaggggct tcatgatgtc cccataattt ttggcagagg gaaaaagatc tcagtggtat
     6961 ttgtgagcca gggcattggc cacaccagcc accaccttct gataggcagc ctgcacctga
     7021 ggagtgcggc cggccgcgat atcctcgacc ccactgggct gtaggtggtg gctggatggc
     7081 ctgagccccc tcctttcctg gggtgcaggc agccacaggg cacaacctgc agtccctggg
     7141 gttggggtag ggagtccgag agcagccaag gggcaggttt ggtgtcatgg gctcagtccc
     7201 cagagcgcac ggggagggga aggttccatg gcaggacctc tgtataggga ctccggggat
     7261 gacgactcgg gctggaggcc aggcaggacg acaggaggtg gggagcagga gaactccggg
     7321 atggggaagc aggagcccca gggacgggac actcagggag cagtggccgt ctccagcagc
     7381 agcagggtcc cggccagcag aggaagcaac gcggggacca cggaccgccc cgggtgcgcg
     7441 gcgtcggtgg tgccggcggg gggcgccagg tcgcaggcgg tgtagggctc caggcaggcg
     7501 gcgaaggcca tgacgtgcgc tatgaaggtc tgctcctgca cgccgtgaac caggtgcgcc
     7561 tgcgggccgc gcgcgaacac cgccacgtcc tcgcctgcgt gggtctcttc gtccaggggc
     7621 actgctgact gctgccgata ctcggggctc ccgctctcgc tctcggtaac atccggccgg
     7681 gcgccgtcct tgagcacata gcctggaccg tttccgtata ggaggaccgt gtaggccttc
     7741 ctgtcccggg ccttgccagg ggccagcccg aagatggagc tccctcgcag ggggtagcct
     7801 ccgaaggaga agacgtggga gtggtcggca gtgacgaggc tcagcgtgtc ctcctcgctg
     7861 gtgagctggc ccgccctctc aatggcgtcg tcgaacatga tcgtctcagt cagtgcccgg
     7921 taagccctgc tttcatgatg accatggtcg atgcgaccac cctccacgaa gaggaagaag
     7981 ccgcgggggt tcctgctcag caggcgcagg gcagcctctg tcatctccat cagggagggg
     8041 tccagtgtgg agtctcggtg gatctcgtat ttcatgtctc caggctcaaa gagacccatg
     8101 agatgggtca cagacgggtc cagggaagcc tgcatgagct cagtgcggtt ccacacatac
     8161 cgggcaccct ggcgcttcgc cagccattcc t
//
