LOCUS       sequence-462841-        5689 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PTCHD4 enhancer, one sensitizing E-box made into buffering 
            Coordinator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5689)
  TITLE     Wysocka Lab enhancer plasmids 2025
REFERENCE   2  (bases 1 to 5689)
  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"
COMMENT     Sequence Label: pGL3_PTCHD4_pos0toneg0
FEATURES             Location/Qualifiers
     source          1..5689
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        17..213
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      64..199
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      243..252
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             249..1901
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(315..333)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    1942..2063
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1953..1972)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2007..2026
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2317..2334)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2488..3076)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2568..2587)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3247..4038,4039..4107))
                     /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     3870..3889
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4108..4212)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4239..4694
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4326..4345)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4536..4557
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4825..4873
                     /note="synthetic polyadenylation signal"
     misc_feature    4887..4978
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4927..4946
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ctcgagatct gcgatctgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
       61 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatcg ctgactaatt
      121 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
      181 ggaggctttt ttggaggcct aggcttttgc aaaaagcttg gcattccggt actgttggta
      241 aagccaccat ggaagacgcc aaaaacataa agaaaggccc ggcgccattc tatccgctgg
      301 aagatggaac cgctggagag caactgcata aggctatgaa gagatacgcc ctggttcctg
      361 gaacaattgc ttttacagat gcacatatcg aggtggacat cacttacgct gagtacttcg
      421 aaatgtccgt tcggttggca gaagctatga aacgatatgg gctgaataca aatcacagaa
      481 tcgtcgtatg cagtgaaaac tctcttcaat tctttatgcc ggtgttgggc gcgttattta
      541 tcggagttgc agttgcgccc gcgaacgaca tttataatga acgtgaattg ctcaacagta
      601 tgggcatttc gcagcctacc gtggtgttcg tttccaaaaa ggggttgcaa aaaattttga
      661 acgtgcaaaa aaagctccca atcatccaaa aaattattat catggattct aaaacggatt
      721 accagggatt tcagtcgatg tacacgttcg tcacatctca tctacctccc ggttttaatg
      781 aatacgattt tgtgccagag tccttcgata gggacaagac aattgcactg atcatgaact
      841 cctctggatc tactggtctg cctaaaggtg tcgctctgcc tcatagaact gcctgcgtga
      901 gattctcgca tgccagagat cctatttttg gcaatcaaat cattccggat actgcgattt
      961 taagtgttgt tccattccat cacggttttg gaatgtttac tacactcgga tatttgatat
     1021 gtggatttcg agtcgtctta atgtatagat ttgaagaaga gctgtttctg aggagccttc
     1081 aggattacaa gattcaaagt gcgctgctgg tgccaaccct attctccttc ttcgccaaaa
     1141 gcactctgat tgacaaatac gatttatcta atttacacga aattgcttct ggtggcgctc
     1201 ccctctctaa ggaagtcggg gaagcggttg ccaagaggtt ccatctgcca ggtatcaggc
     1261 aaggatatgg gctcactgag actacatcag ctattctgat tacacccgag ggggatgata
     1321 aaccgggcgc ggtcggtaaa gttgttccat tttttgaagc gaaggttgtg gatctggata
     1381 ccgggaaaac gctgggcgtt aatcaaagag gcgaactgtg tgtgagaggt cctatgatta
     1441 tgtccggtta tgtaaacaat ccggaagcga ccaacgcctt gattgacaag gatggatggc
     1501 tacattctgg agacatagct tactgggacg aagacgaaca cttcttcatc gttgaccgcc
     1561 tgaagtctct gattaagtac aaaggctatc aggtggctcc cgctgaattg gaatccatct
     1621 tgctccaaca ccccaacatc ttcgacgcag gtgtcgcagg tcttcccgac gatgacgccg
     1681 gtgaacttcc cgccgccgtt gttgttttgg agcacggaaa gacgatgacg gaaaaagaga
     1741 tcgtggatta cgtcgccagt caagtaacaa ccgcgaaaaa gttgcgcgga ggagttgtgt
     1801 ttgtggacga agtaccgaaa ggtcttaccg gaaaactcga cgcaagaaaa atcagagaga
     1861 tcctcataaa ggccaagaag ggcggaaaga tcgccgtgta attctagagt cggggcggcc
     1921 ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     1981 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     2041 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     2101 agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcg
     2161 ataaggatct ggatccgtcg accgatgccc ttgagagcct tcaacccagt cagctccttc
     2221 cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa
     2281 ctcgtaggac aggtgccggc agcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     2341 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     2401 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     2461 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     2521 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     2581 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     2641 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     2701 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     2761 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     2821 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     2881 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     2941 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     3001 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     3061 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     3121 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     3181 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     3241 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     3301 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     3361 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     3421 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     3481 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     3541 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     3601 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     3661 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     3721 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     3781 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     3841 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     3901 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     3961 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     4021 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     4081 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     4141 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     4201 aggggttccg cgcacatttc cccgaaaagt gccacctgac gcgccctgta gcggcgcatt
     4261 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     4321 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     4381 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     4441 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     4501 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     4561 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     4621 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     4681 aacgcttaca atttgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
     4741 gcgggcctct tcgctattac gccagcccaa gctaccatga taagtaagta atattaaggt
     4801 acgggaggta cttggagcgg ccgcaataaa atatctttat tttcattaca tctgtgtgtt
     4861 ggttttttgt gtgaatcgat agtactaaca tacgctctcc atcaaaacaa aacgaaacaa
     4921 aacaaactag caaaataggc tgtccccagt gcaagtgcag gtgccagaac atttctctat
     4981 cgataggtac cgagctctta cgcgtgctag cccttgctca accttatact cagcttaatt
     5041 aaaaccctgc tattactaat tgagaaacta tcacttattt gttcaaacta ccctaactga
     5101 gccctctatt aatttgttag taagaacact agcatctgga gtagtatctc atagctgttg
     5161 gttgcactta gagaagaatc ttatttttta aattttctct acttttattc taggttcagg
     5221 gagtacatgt gcagatttgt tacatgggtc aattttgtgt cactaaggtt tggtgtacaa
     5281 atgaaagcat agaaccgaat agtttttcaa ccgatgtttc cctcccaccc tcccatgtca
     5341 atcagtccac agtgtctatc attcccatct ttgtgtccac atgtattcaa tgtttagctc
     5401 ctgattataa gtgagaacat gtggtttcta ttattctctt cctgcattag tttgcttagc
     5461 ataatagctt ccagctacct ccatgttgct tgcaaaggac atgattgtgt tactttgtgg
     5521 gagaatcttt aaagttaaga aagtatttaa aatttcttca agatttaaaa ttctgctatg
     5581 tttcaagatt aaaacattta agcactttga gtatgtgtga atacatgtgc atattttgat
     5641 ttttcatatt atttcatatt tcatacgtat tctgagagtc caattgtga
//
