LOCUS       sequence-427345-        5858 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  transfer plasmid for AAV vector production of mCherry and miR for Mm
            ATP10B.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5858)
  AUTHORS   Sanchiz-Calvo M, Coccia E, Cawthorne C, Morrone Parfitt G, 
            Torre-Muruzabal T, Tsafaras G, Van Laere K, Cabezudo D, Cascalho A, 
            Van den Haute C, Vangheluwe P, Blanchard J, Bentea E, Baekelandt V
  TITLE     Loss of the lysosomal lipid flippase ATP10B leads to progressive 
            dopaminergic neurodegeneration and parkinsonian motor deficits.
  JOURNAL   Acta Neuropathol. 2025 Jul 17;150(1):5. doi: 
            10.1007/s00401-025-02908-0.
  PUBMED    40676227
REFERENCE   2  (bases 1 to 5858)
  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: "Acta 
            Neuropathol."; date: "2025-07-17"; volume: "150(1):5. doi"; pages: "
            10.1007/s00401-025-02908-0"
COMMENT     Sequence Label: pAAV-CMVenh synapsin-intron-mCherry-1x miR5 Mm...
FEATURES             Location/Qualifiers
     source          1..5858
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(136..153)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(136..152)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(145..167)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      294..749
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(381..400)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     591..612
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        775..879
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(880..948,949..1740)
                     /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(1098..1117)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1911..2499
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2400..2419
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    2787..2808
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2823..2828,2829..2846,2847..2853)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2861..2877
                     /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     2866..2888
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2885..2901
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2885..2901
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   2939..3068
                     /label=AAV2 ITR
     enhancer        3209..3512
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          3947..4079
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     4062..4081
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      4116..4125
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4122..4832
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4267..4285)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4535..4554
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     ncRNA           4897..4911
                     /label=miR-30a loop
                     /note="loop from the the 71-nt precursor of the human 
                     miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     primer_bind     complement(5049..5069)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5467..5478)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5545..5562)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5551..5758
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   join(5846..5858,1..117)
                     /label=AAV2 ITR
ORIGIN
        1 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
       61 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagcct
      121 taattaacct aattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt
      181 tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga
      241 ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc
      301 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
      361 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
      421 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
      481 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
      541 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
      601 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
      661 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
      721 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
      781 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
      841 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
      901 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
      961 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     1021 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     1081 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     1141 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     1201 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     1261 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     1321 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     1381 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     1441 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     1501 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     1561 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     1621 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     1681 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     1741 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     1801 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     1861 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     1921 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     1981 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     2041 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     2101 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2161 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2221 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2281 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2341 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2401 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2461 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2521 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     2581 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     2641 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
     2701 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     2761 aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag
     2821 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt
     2881 cacacaggaa acagctatga ccatgattac gccagattta attaaggcct taattaggct
     2941 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg
     3001 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     3061 gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc catgctctag
     3121 gaagatcgga attcgccctt aagctagata gttattaata gtaatcaatt acggggtcat
     3181 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg
     3241 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa
     3301 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
     3361 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
     3421 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt
     3481 acatctacgt attagtcatc gctattacca tggcacctgc agagggccct gcgtatgagt
     3541 gcaagtgggt tttaggacca ggatgaggcg gggtgggggt gcctacctga cgaccgaccc
     3601 cgacccactg gacaagcacc caacccccat tccccaaatt gcgcatcccc tatcagagag
     3661 ggggagggga aacaggatgc ggcgaggcgc gtgcgcactg ccagcttcag caccgcggac
     3721 agtgccttcg cccccgcctg gcggcgcgcg ccaccgccgc ctcagcactg aaggcgcgct
     3781 gacgtcactc gccggtcccc cgcaaactcc ccttcccggc caccttggtc gcgtccgcgc
     3841 cgccgccggc ccagccggac cgcaccacgc gaggcgcgag ataggggggc acgggcgcga
     3901 ccatctgcgc tgcggccgca ctagttggtc gtgaggcact gggcaggtaa gtatcaaggt
     3961 tacaagacag gtttaaggag accaatagaa actgggcttg tcgagacaga gaagactctt
     4021 gcgtttctga taggcaccta ttggtcttac tgacatccac tttgcctttc tctccacagg
     4081 tgtccaggcg gccgctctag aggatcccca ccggtgccac catggtgagc aagggcgagg
     4141 aggataacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg
     4201 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc
     4261 agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttcgcctgg gacatcctgt
     4321 cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac atccccgact
     4381 acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg
     4441 gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggcgagttc atctacaagg
     4501 tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag aagaccatgg
     4561 gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag ggcgagatca
     4621 agcagaggct gaagctgaag gacggcggcc actacgacgc tgaggtcaag accacctaca
     4681 aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag ttggacatca
     4741 cctcccacaa cgaggactac accatcgtgg aacagtacga acgcgccgag ggccgccact
     4801 ccaccggcgg catggacgag ctgtacaagt aactcgagat ccaagaaggt atattgctgt
     4861 tgacagtgag cgacctggtg attctgaact ggattagtga agccacagat gtaatccagt
     4921 tcagaatcac caggatgcct actgcctcgg acttcaaggg ctacgatgtc gacttggatc
     4981 catcgcgcgt gatccaatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     5041 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     5101 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     5161 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     5221 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     5281 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     5341 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
     5401 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
     5461 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
     5521 cctcttccgc gtcttcgaga tctgcctcga ctgtgccttc tagttgccag ccatctgttg
     5581 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     5641 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     5701 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggact
     5761 cgagttaagg gcgaattccc gataaggatc ttcctagagc atggctacgt agataagtag
     5821 catggcgggt taatcattaa ctacaaggaa cccctagt
//
