LOCUS       sequence-429604-        5983 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector to drive the expression of eGFP under the control of the 
            4xBRE regulatory element of SMAD1 and miniXon splicing casette 
            transcription factor in vivo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5983)
  AUTHORS   Okur Z, Schlauri N, Bitsikas V, Panopoulou M, Karmakar K, Schreiner 
            D, Scheiffele P
  TITLE     Control of neuronal excitation-inhibition balance by BMP-SMAD1 
            signaling
  JOURNAL   bioRxiv 2023
REFERENCE   2  (bases 1 to 5983)
  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: "bioRxiv 
            2023"
COMMENT     Sequence Label: pAAV-BRX-eGFP-NLS
FEATURES             Location/Qualifiers
     source          1..5983
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   651..780
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     polyA_signal    813..861
                     /note="synthetic polyadenylation signal"
     misc_feature    875..966
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     915..934
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        1084..1280
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1131..1266
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      1823..1832
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1926..1928,1929..2639)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1926..1928,1929..2639)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1968..1989)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2229..2248)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2576..2597
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2649..2669
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2673..2693
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2697..2717
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    2760..3348
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2813..2833)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3231..3242)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3380..3856
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3579..3598)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3896..4036
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3896..4025
                     /label=AAV2 ITR
     rep_origin      4111..4566
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4198..4217)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4408..4429
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4583..4602)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4702..4724
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4762..4780)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4848..4952
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4953..5021,5022..5813)
                     /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(5171..5190)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt cctgcaggca
      661 gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt ggtcgcccgg
      721 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      781 gcggccgcac gcgccgtact tggagcggcc gcaataaaat atctttattt tcattacatc
      841 tgtgtgttgg ttttttgtgt gaatcgatag tactaacata cgctctccat caaaacaaaa
      901 cgaaacaaaa caaactagca aaataggctg tccccagtgc aagtgcaggt gccagaacat
      961 ttctctatcg ataggtaccg ggcctggcgt ctaacggtct gagggcctgg cgtctaacgg
     1021 tctgagggcc tggcgtctaa cggtctgagg gcctggcgtc taacggtctg aagatctgcg
     1081 atctgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     1141 taactccgcc cagttccgcc cattctccgc cccatcgctg actaattttt tttatttatg
     1201 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
     1261 gaggcctagg cttttgcaaa aagcttggca ttccggtagc tggtcgactt atacgaagtt
     1321 atgctagctt ggcaatccgg tactgttggt aaatttctgt acaacttaac cttgcagaga
     1381 gccactggca tcagctttgc cattcttgga aacttttctg gtaagttctc tcgttaccat
     1441 cttttgaaat tttaagtgaa ttaatacata tcttgcttag tctcttgtgc aggaaatgtt
     1501 ttccatttat gacaaaacag gaattgtgtg aaatttcaaa tattggatta ggaaatacaa
     1561 agttactgaa agtgaggtac taatgtttat aaaataaaaa ctttttcttg ccatttgcag
     1621 atttaacatt tttgagtcaa tccaagtgcc accatgcagg aggttcatga ttgtgtagag
     1681 taagacataa ttttgttgag gtttaactct gaatacttaa tgtggtactg aatacttaat
     1741 gtggtactga gaggcagcct aactgaccac acagcattca cgcctggcta atttttgtat
     1801 ttttagtaga gacggggttt caccatggtg gccaggctgg gttctggttg tttatgatct
     1861 ttattttttg gtgatctagg aaccaaacaa caagaaattg ttgtttcccg tgggtaagag
     1921 gatctgtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     1981 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     2041 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     2101 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     2161 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     2221 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     2281 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     2341 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     2401 acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     2461 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     2521 actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     2581 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg
     2641 gatccgatcc aaaaaagaag agaaaggtag atccaaaaaa gaagagaaag gtagatccaa
     2701 aaaagaagag aaaggtatga tctagagggc cctatgaatt cgatatcaag cttatcgata
     2761 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     2821 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     2881 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     2941 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     3001 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     3061 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     3121 tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg
     3181 cctatgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     3241 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     3301 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgag
     3361 cgctgctcga gagatctacg ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc
     3421 cctggaagtt gccactccag tgcccaccag ccttgtccta ataaaattaa gttgcatcat
     3481 tttgtctgac taggtgtcct tctataatat tatggggtgg aggggggtgg tatggagcaa
     3541 ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc tattgggaac caagctggag
     3601 tgcagtggca caatcttggc tcactgcaat ctccgcctcc tgggttcaag cgattctcct
     3661 gcctcagcct cccgagttgt tgggattcca ggcatgcatg accaggctca gctaattttt
     3721 gtttttttgg tagagacggg gtttcaccat attggccagg ctggtctcca actcctaatc
     3781 tcaggtgatc tacccacctt ggcctcccaa attgctggga ttacaggcgt gaaccactgc
     3841 tcccttccct gtccttctga ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa
     3901 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     3961 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     4021 cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     4081 tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg
     4141 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc
     4201 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     4261 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     4321 ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     4381 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     4441 actctatctc gggctattct tttgatttat aagggatttt gccgatttcg gtctattggt
     4501 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta
     4561 caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     4621 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     4681 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     4741 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     4801 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     4861 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     4921 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     4981 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     5041 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     5101 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     5161 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     5221 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     5281 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     5341 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     5401 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     5461 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     5521 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     5581 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     5641 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     5701 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     5761 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     5821 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     5881 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     5941 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
