LOCUS       Exported                7276 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV vector with hSynapsin promoter, Cre-OFF mCAR (for efficient 
            CAV-2 infection), Cre-ON DTR-EGFP (diphtheria toxin receptor for 
            cell ablation).
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-hSyn-DIO {mCAR}off-{DTR-GFP}on-WPRE
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7276)
  AUTHORS   Li SJ, Vaughan A, Sturgill JF, Kepecs A
  TITLE     A Viral Receptor Complementation Strategy to Overcome CAV-2 Tropism 
            for Efficient Retrograde Targeting of Neurons.
  JOURNAL   Neuron. 2018 Jun 6;98(5):905-917.e5. doi: 
            10.1016/j.neuron.2018.05.028.
  PUBMED    29879392
REFERENCE   2  (bases 1 to 7276)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7276
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        175..622
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    660..693
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    744..777
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     regulatory      783..792
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1845..1874
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(1886..2605)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(1889..2605)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1931..1952)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2280..2299
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2539..2560
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2602..2611
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     protein_bind    complement(3261..3294)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     protein_bind    complement(3345..3378)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    3403..3991
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3456..3476)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3874..3885)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4023..4499
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4222..4241)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4539..4679
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4539..4668
                     /label=AAV2 ITR
     rep_origin      4754..5209
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4841..4860)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5051..5072
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5226..5245)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5345..5367
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5405..5423)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5491..5595
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5596..6456
                     /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(5814..5833)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6627..7215
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7116..7135
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtgtgtct agactgcaga gggccctgcg tatgagtgca
      181 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      241 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      301 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      361 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      421 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      481 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      541 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      601 cgtgtcgtgc ctgagagcgc agtcgagaag gtaccggatc ctctagagtc gactccggaa
      661 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
      721 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
      781 gcgccaccat ggctcgccta ctgtgcttcg tgctcttgtg cgggatcgcg gatttcacca
      841 gtggtttgag catcactaca cccgaacaga ggatcgaaaa agccaaaggg gaaactgcgt
      901 atctaccatg caagtttact ctcagtcccg aagaccaggg accactggac attgaatggc
      961 tgatatcccc gtctgataac cagatagtgg atcaagtgat cattttgtat tctggagaca
     1021 aaatttatga taactactat ccggatctga aaggacgggt acattttacg agtaacgatg
     1081 tcaagtctgg cgacgcatct ataaatgtga ccaacctgca gctgtcggac attggcactt
     1141 accagtgcaa agtgaagaaa gcccctgggg ttgcaaataa gaaattcctg ctgaccgttc
     1201 ttgttaagcc ttcaggtaca agatgcttcg tggatggatc ggaagagatt ggaaatgact
     1261 tcaagctaaa atgtgaaccc aaggaaggct cccttccact acagtttgaa tggcagaaac
     1321 tgtcggactc ccagacaatg cctacgccat ggctggcaga aatgacgtca ccagttatat
     1381 ctgtgaagaa cgccagttct gagtattctg ggacatacag ctgcacggtt caaaacagag
     1441 tgggctctga ccagtgtatg ctgcgactag acgttgtccc accctccaac cgagccggaa
     1501 cgatcgcggg cgccgtcata gggacgctgc tggcccttgt gctcatcggg gccatcctct
     1561 tctgctgtca caggaaacgc agagaagaga agtacgagaa ggaagttcat catgatatca
     1621 gggaagatgt gcctcctcca aagagtcgga catccactgc caggagctat attggcagca
     1681 accattcctc cctgggatcc atgtccccct ctaacatgga ggggtattcc aagacgcagt
     1741 ataaccaagt ccccagtgaa gactttgaac gtgcgcctca gagcccgact ctggcacccg
     1801 ctaagttcaa gtacgcttac aagaccgatg gcattacagt ggtagagcag aagctgatca
     1861 gcgaggagga cctgtaatta attaattact tgtacagctc gtccatgccg agagtgatcc
     1921 cggcggcggt cacgaactcc agcaggacca tgtgatcgcg cttctcgttg gggtctttgc
     1981 tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga
     2041 tgggggtgtt ctgctggtag tggtcggcga gctgcacgct gccgtcctcg atgttgtggc
     2101 ggatcttgaa gttcaccttg atgccgttct tctgcttgtc ggccatgata tagacgttgt
     2161 ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga
     2221 tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg
     2281 tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca
     2341 tggcggactt gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg aagcactgca
     2401 cgccgtaggt cagggtggtc acgagggtgg gccagggcac gggcagcttg ccggtggtgc
     2461 agatgaactt cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc
     2521 tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag gatgggcacc accccggtga
     2581 acagctcctc gcccttgctc accatggtgg cgaccggtgg atccgggtgg gaattagtca
     2641 tgcccaactt cactttctct tcgttttcca catcataacc acctctccta tggtacctaa
     2701 acatgagaag ccccacgatg accagcagac agacagagga cagcaccacg gccaccacag
     2761 ccaggatagt tgtatggtca taggtatata agcgattttc cactgggagg ctcagcccat
     2821 gacacctctc tccatggtaa cctgggtggc agatgcagga gggagcccgg agctccttca
     2881 catatttgca ttctccgtgg atgcagaagt ccttgtattt ccgaagacat gggtccctct
     2941 tcttccctag tcccttgcct ttcttctttc ttttcccgtg ctcctccttg cttggtgtgg
     3001 ccagtgcttg tggcttggag gataaagtga ctctcaaaag gtccagatct gcctcttgca
     3061 agtcacggac tttccggtcc cggccgcctc ctaggcgtag cagctggtcc gtagatccag
     3121 tggaagggtc cgggttgctg gttccagcag ctagccctct ccgaagctgc tccaggctct
     3181 cgccagtcac cagtgccgaa agaactgcag ccagaaggag cttcagcacc accgacggca
     3241 gcagcttcat ggtgactagc ataacttcgt ataaagtatc ctatacgaag ttatttgcct
     3301 taacccagaa attatcactg ttattcttta gaatggtgca aagaataact tcgtataatg
     3361 tatgctatac gaagttatga attcgatatc aagcttatcg ataatcaacc tctggattac
     3421 aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga
     3481 tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc
     3541 tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa
     3601 cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc
     3661 acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc
     3721 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc
     3781 gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctatgt tgccacctgg
     3841 attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct
     3901 tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg
     3961 agtcggatct ccctttgggc cgcctccccg catcgatacc gagcgctgct cgagagatct
     4021 acgggtggca tccctgtgac ccctccccag tgcctctcct ggccctggaa gttgccactc
     4081 cagtgcccac cagccttgtc ctaataaaat taagttgcat cattttgtct gactaggtgt
     4141 ccttctataa tattatgggg tggagggggg tggtatggag caaggggcaa gttgggaaga
     4201 caacctgtag ggcctgcggg gtctattggg aaccaagctg gagtgcagtg gcacaatctt
     4261 ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt
     4321 tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac
     4381 ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg atctacccac
     4441 cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc
     4501 tgattttgta ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt
     4561 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     4621 acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg
     4681 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca
     4741 aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     4801 cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct
     4861 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     4921 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt
     4981 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     5041 ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat
     5101 tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt
     5161 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact
     5221 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     5281 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     5341 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     5401 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     5461 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     5521 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     5581 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     5641 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5701 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5761 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     5821 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     5881 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     5941 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     6001 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     6061 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     6121 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     6181 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     6241 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     6301 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     6361 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     6421 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     6481 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     6541 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     6601 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     6661 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     6721 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     6781 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     6841 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     6901 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     6961 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     7021 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     7081 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     7141 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     7201 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     7261 ccttttgctc acatgt
//
