LOCUS       sequence-273099-        5924 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Minigene reporter for alternative splicing analysis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5924)
  AUTHORS   Zheng S, Damoiseaux R, Chen L, Black DL
  TITLE     A broadly applicable high-throughput screening strategy identifies 
            new regulators of Dlg4 (Psd-95) alternative splicing.
  JOURNAL   Genome Res. 2013 Jun;23(6):998-1007. doi: 10.1101/gr.147546.112. 
            Epub 2013 May 1.
  PUBMED    23636947
REFERENCE   2  (bases 1 to 5924)
  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: 
            "10.1101/gr.147546.112"; journalName: "Genome Res"; date: "2013-06";
            volume: "23"; issue: "6"; pages: "998-1007"
COMMENT     Sequence Label: pFlare5GdP
FEATURES             Location/Qualifiers
     source          1..5924
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        66..369
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        370..573
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     524..544
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1011..1020
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1156..1158,1159..1872)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHLKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGTDELYK"
     primer_bind     complement(1198..1219)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1459..1478)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     regulatory      1879..1888
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1885..2595
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent 
                     protein"
                     /label=mStrawberry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILTPNFTYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKMRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYIVGIKLDITSHNEDYTIVELYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2030..2048)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2298..2317
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2717..2838
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2754..2773)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2808..2827
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2845..3300)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2982..3003)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3194..3213
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3327..3431
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3433..3790
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3433..3624
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3456..3476)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3641..3776
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3825..4619
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3825..4619
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3879..3898)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4489..4508
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4807..4826)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4851..4898
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5227..5815
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5716..5735
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
       61 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
      121 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
      181 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      241 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      301 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      361 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
      421 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
      481 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
      541 cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca gatccgctag
      601 cgctaccgga ctcagatcta ccattggtgc acctgactcc tgaggagaag tctgccgtta
      661 ctgccctgtg gggcaaggtg aacgtggaag agttggtggt gaggccctgg gccacaggta
      721 agtatcaagg ttacaagaca ggtttaagga gaccaataga aactgggcat gtggagacag
      781 agaagactct tgggtttctg aattcattgg atctaccatt ggtgcacctg actcctgagg
      841 agaagtctgc cgttactgcc ctgtggggca aggtgaacgt ggaagagttg gtggtgaggc
      901 cctgggccac aggtaagtat caaggttaca agacaggttt aaggagacca atagaaactg
      961 ggcatgtgga gacagagaag actcttgggt ttctgaattc attggatcca ccatggtggc
     1021 ttagatccgg gcatgtggag acagagaaga ctgttgagtt tgtgataagc actgactctc
     1081 tctgcctatt ggtctatttt ccctccctca gtggcggtcg aggacaaact ctacgcggac
     1141 ttgccagtac gcttcgtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     1201 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     1261 gacgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     1321 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgctttag ccgctacccc
     1381 gaccacctga agcagcacga cttcttcaag agtgcaatgc ccgaaggcta cgtccaggag
     1441 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     1501 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1561 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat caccgccgac
     1621 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1681 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1741 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1801 gatcatatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg caccgacgag
     1861 ctgtacaagt aaccggtcgc caccatggtg agcaagggcg aggagaataa catggccatc
     1921 atcaaggagt tcatgcgctt caaggtgcgc atggagggct ccgtgaacgg ccacgagttc
     1981 gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag
     2041 gtgaccaagg gtggccccct gcccttcgcc tgggacatcc taacccccaa cttcacctac
     2101 ggctccaagg cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc
     2161 cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg
     2221 acccaggact cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc
     2281 aacttcccct ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc
     2341 gagcggatgt accccgagga cggcgccctg aagggcgaga tcaagatgag gctgaagctg
     2401 aaggacggcg gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg
     2461 cagctgcccg gcgcctacat cgtcggcatc aagttggaca tcacctccca caacgaggac
     2521 tacaccatcg tggaactgta cgaacgcgcc gagggccgcc actccaccgg cggcatggac
     2581 gagctgtata agtaagcggc cgcgactcta gatcataatc agccatacca catttgtaga
     2641 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     2701 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     2761 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     2821 actcatcaat gtatcttaag gcgtaaattg taagcgttaa tattttgtta aaattcgcgt
     2881 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     2941 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     3001 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     3061 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     3121 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     3181 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     3241 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     3301 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     3361 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     3421 aaaggaagag tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg
     3481 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     3541 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3601 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     3661 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     3721 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     3781 cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     3841 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     3901 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     3961 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc
     4021 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     4081 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     4141 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     4201 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     4261 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     4321 cgccagccga actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg
     4381 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     4441 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     4501 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     4561 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     4621 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     4681 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     4741 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag
     4801 gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa
     4861 aagacagaat aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     4921 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     4981 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     5041 cggggcggca ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa
     5101 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     5161 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     5221 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     5281 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     5341 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     5401 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5461 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5521 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5581 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5641 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5701 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5761 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5821 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     5881 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcca
//
