LOCUS       sequence-235533-        5361 bp    DNA     circular SYN 15-APR-2025
DEFINITION  pAAV plasmid expressing mCherry.3XFLAG under the hSyn promoter with 
            a Linker containing BstBI, BspQI and SacI sites upstream of the hSyn
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5361)
  AUTHORS   Grunewald L, Chiocchetti AG, Weber H, Scholz CJ, Schartner C, 
            Freudenberg F, Reif A
  TITLE     Knockdown of the ADHD Candidate Gene Diras2 in Murine Hippocampal 
            Primary Cells.
  JOURNAL   J Atten Disord. 2019 Jan 9:1087054718822129. doi: 
            10.1177/1087054718822129.
  PUBMED    30623719
REFERENCE   2  (bases 1 to 5361)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Atten 
            Disord. 2019 Jan 9:1087054718822129. doi: 10.1177/1087054718822129."
COMMENT     Sequence Label: pAAV-Linker-hSyn::mCherry.3xFLAG-WPRE
FEATURES             Location/Qualifiers
     source          1..5361
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      114..569
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(201..220)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     411..432
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(586..605)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     705..727
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(765..783)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        851..955
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(956..1024,1025..1816)
                     /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(1174..1193)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1987..2575
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2476..2495
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   2637..2766
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        2840..3287
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      3319..3328
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3325..4032
                     /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(3470..3488)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3738..3757
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             4039..4104
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    4124..4712
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4177..4197)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4595..4606)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4744..5220
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4943..4962)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   join(5260..5361,1..39)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   join(5260..5361,1..28)
                     /label=AAV2 ITR
ORIGIN
        1 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc
       61 tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc
      121 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
      181 tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
      241 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
      301 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
      361 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
      421 gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat
      481 tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
      541 ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga
      601 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
      661 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
      721 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
      781 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
      841 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
      901 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
      961 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     1021 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     1081 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     1141 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     1201 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     1261 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     1321 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     1381 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     1441 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     1501 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     1561 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     1621 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     1681 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     1741 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     1801 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     1861 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     1921 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     1981 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2041 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2101 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     2161 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     2221 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     2281 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     2341 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     2401 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     2461 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     2521 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     2581 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtcctg
     2641 caggcagctg cgcgctcgct cgctcactga ggccgcccgg gcgtcgggcg acctttggtc
     2701 gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg
     2761 gttcctgcgg ccgcacgcgt tcgaacgagc tcgctcttcg ttttcgcgtg tgtctagact
     2821 gcagagggcc ctgcgtatga gtgcaagtgg gttttaggac caggatgagg cggggtgggg
     2881 gtgcctacct gacgaccgac cccgacccac tggacaagca cccaaccccc attccccaaa
     2941 ttgcgcatcc cctatcagag agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac
     3001 tgccagcttc agcaccgcgg acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc
     3061 gcctcagcac tgaaggcgcg ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg
     3121 gccaccttgg tcgcgtccgc gccgccgccg gcccagccgg accgcaccac gcgaggcgcg
     3181 agataggggg gcacgggcgc gaccatctgc gctgcggcgc cggcgactca gcgctgcctc
     3241 agtctgcggt gggcagcgga ggagtcgtgt cgtgcctgag agcgcagtcg agaaggtacc
     3301 ggatcctcta gagtcgacgc caccatggtg agcaagggcg aggaggataa catggccatc
     3361 atcaaggagt tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc
     3421 gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag
     3481 gtgaccaagg gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac
     3541 ggctccaagg cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc
     3601 cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg
     3661 acccaggact cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc
     3721 aacttcccct ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc
     3781 gagcggatgt accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg
     3841 aaggacggcg gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg
     3901 cagctgcccg gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac
     3961 tacaccatcg tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac
     4021 gagctgtaca aggaattcga ctacaaagac catgacggtg attataaaga tcatgatatt
     4081 gattacaagg atgacgatga caagctagca taagcttatc gataatcaac ctctggatta
     4141 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     4201 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     4261 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     4321 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     4381 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     4441 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     4501 cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg
     4561 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     4621 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     4681 gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc
     4741 tacgggtggc atccctgtga cccctcccca gtgcctctcc tggccctgga agttgccact
     4801 ccagtgccca ccagccttgt cctaataaaa ttaagttgca tcattttgtc tgactaggtg
     4861 tccttctata atattatggg gtggaggggg gtggtatgga gcaaggggca agttgggaag
     4921 acaacctgta gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt ggcacaatct
     4981 tggctcactg caatctccgc ctcctgggtt caagcgattc tcctgcctca gcctcccgag
     5041 ttgttgggat tccaggcatg catgaccagg ctcagctaat ttttgttttt ttggtagaga
     5101 cggggtttca ccatattggc caggctggtc tccaactcct aatctcaggt gatctaccca
     5161 ccttggcctc ccaaattgct gggattacag gcgtgaacca ctgctccctt ccctgtcctt
     5221 ctgattttgt aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt
     5281 tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc
     5341 gacgcccggg ctttgcccgg g
//
