LOCUS       Exported                7422 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV vector with hSynapsin promoter, Cre-OFF hCAR (for efficient 
            CAV-2 infection), Cre-ON hM4Di-mCherry (Gi-coupled DREADD for 
            neuronal silencing), W3SL cassette (for maximize cloning capacity).
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-hSyn-DIO {hCAR}off-{hM4Di-mCherry}on-W3SL
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7422)
  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 7422)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7422
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        186..633
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    671..704
                     /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    755..788
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     regulatory      794..803
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1895..1924
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(1936..2646)
                     /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(2214..2233)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2483..2501
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      2643..2652
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     regulatory      4101..4110
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     protein_bind    complement(4117..4150)
                     /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(4201..4234)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     complement(4304..4324)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4684..4824
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4684..4813
                     /label=AAV2 ITR
     rep_origin      4899..5354
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4986..5005)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5196..5217
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5371..5390)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5490..5512
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5550..5568)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5636..5740
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5741..6601
                     /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(5959..5978)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6772..7360
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7261..7280
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtgtgtc tagactgcag agggccctgc
      181 gtatgagtgc aagtgggttt taggaccagg atgaggcggg gtgggggtgc ctacctgacg
      241 accgaccccg acccactgga caagcaccca acccccattc cccaaattgc gcatccccta
      301 tcagagaggg ggaggggaaa caggatgcgg cgaggcgcgt gcgcactgcc agcttcagca
      361 ccgcggacag tgccttcgcc cccgcctggc ggcgcgcgcc accgccgcct cagcactgaa
      421 ggcgcgctga cgtcactcgc cggtcccccg caaactcccc ttcccggcca ccttggtcgc
      481 gtccgcgccg ccgccggccc agccggaccg caccacgcga ggcgcgagat aggggggcac
      541 gggcgcgacc atctgcgctg cggcgccggc gactcagcgc tgcctcagtc tgcggtgggc
      601 agcggaggag tcgtgtcgtg cctgagagcg cagtcgagaa ggtaccggat cctctagagt
      661 cgactccgga ataacttcgt ataggatact ttatacgaag ttatgcagaa tggtagctgg
      721 attgtagctg ctattagcaa tatgaaacct cttaataact tcgtatagca tacattatac
      781 gaagttatgg cgcgccacca tggcgctcct gctgtgcttc gtgctcctgt gcggagtagt
      841 ggatttcgcc agaagtttga gtatcactac tcctgaagag atgattgaaa aagccaaagg
      901 ggaaactgcc tatctgccgt gcaaatttac gcttagtccc gaagaccagg gaccgctgga
      961 catcgagtgg ctgatatcac cagctgataa tcagaaggtg gatcaagtga ttattttata
     1021 ttctggagac aaaatttatg atgactacta tccagatctg aaaggccgag tacattttac
     1081 gagtaatgat ctcaaatctg gtgatgcatc aataaatgta acgaatttac aactgtcaga
     1141 tattggcaca tatcagtgca aagtgaaaaa agctcctggt gttgcaaata agaagattca
     1201 tctggtagtt cttgttaagc cttcaggtgc gagatgttac gttgatggat ctgaagaaat
     1261 tggaagtgac tttaagataa aatgtgaacc aaaagaaggt tcacttccat tacagtatga
     1321 gtggcaaaaa ttgtctgact cacagaaaat gcccacttca tggttagcag aaatgacttc
     1381 atctgttata tctgtaaaaa atgcctcttc tgagtactct gggacataca gctgtacagt
     1441 cagaaacaga gtgggctctg atcagtgcct gttgcgtcta aacgttgtcc ctccttcaaa
     1501 taaagctgga ctaattgcag gagccattat aggaactttg cttgctctag cgctcattgg
     1561 tcttatcatc ttttgctgtc gtaaaaagcg cagagaagaa aaatatgaaa aggaagttca
     1621 tcacgatatc agggaagatg tgccacctcc aaagagccgt acgtccactg ccagaagcta
     1681 catcggcagt aatcattcat ccctggggtc catgtctcct tccaacatgg aaggatattc
     1741 caagactcag tataaccaag taccaagtga agactttgaa cgcactcctc agagtccgac
     1801 tctcccacct gctaaggtag ctgcccctaa tctaagtcga atgggtgcga ttcctgtgat
     1861 gattccagca cagagcaagg atgggtctat agtagagcag aagctgatca gcgaggagga
     1921 cctgtaatta attaattact tgtacagctc gtccatgccg ccggtggagt ggcggccctc
     1981 ggcgcgttcg tactgttcca cgatggtgta gtcctcgttg tgggaggtga tgtccaactt
     2041 gatgttgacg ttgtaggcgc cgggcagctg cacgggcttc ttggccttgt aggtggtctt
     2101 gacctcagcg tcgtagtggc cgccgtcctt cagcttcagc ctctgcttga tctcgccctt
     2161 cagggcgccg tcctcggggt acatccgctc ggaggaggcc tcccagccca tggtcttctt
     2221 ctgcattacg gggccgtcgg aggggaagtt ggtgccgcgc agcttcacct tgtagatgaa
     2281 ctcgccgtcc tgcagggagg agtcctgggt cacggtcacc acgccgccgt cctcgaagtt
     2341 catcacgcgc tcccacttga agccctcggg gaaggacagc ttcaagtagt cggggatgtc
     2401 ggcggggtgc ttcacgtagg ccttggagcc gtacatgaac tgaggggaca ggatgtccca
     2461 ggcgaagggc agggggccac ccttggtcac cttcagcttg gcggtctggg tgccctcgta
     2521 ggggcggccc tcgccctcgc cctcgatctc gaactcgtgg ccgttcacgg agccctccat
     2581 gtgcaccttg aagcgcatga actccttgat gatggccatg ttatcctcct cgcccttgct
     2641 caccatggtg gcgaccggtg gatcccgcct ggcagtgccg atgttccgat actggcacag
     2701 cagcaggtgc cggaaggtct ttttaaaggt ggcgttgcac agagcatagc aggcagggtt
     2761 gatggtgctg ttgacgtagc agagccagta gccaatggac cacaccgtgt cagggatgca
     2821 gctctggcag aaggtgttca ccaggaccat gacgttgtag ggcgtccagg tgaggatgaa
     2881 ggccagcaga atggcaaaga tcgttcgtgt cactttgcgc tcccgggccg ccatctgccg
     2941 cttcttgcgc acctggttgc gagcgatgct ggcgaacttg cgggccacgt tggccgcagg
     3001 gcgcatgcca gccggcgtgg caggcacaat ctcaatggct gtcacacact cattgcctgt
     3061 ctgcttcgtc acaatctgga tcttggacca tctggaggct gggttgaggg cccgcggctg
     3121 caggggaggg gcgggcatgg cgggcgtggt ggcctctgtg gtggacagct ctgtggctgg
     3181 gcgttccttg gtgttctggg tggcactgcc tgagctggac tcattggaag tgtccttatc
     3241 agccacgggg cgcggtggcg gtggcagcgc tggcgggggg gcctcctcca gcttgccatt
     3301 gcgcagctcc tcccgggcgg cctccccggg cgggggcttc ttgacgctct gcttcattag
     3361 tgggctcttg aggaaggcca gcgtcttggc tttcttctcc ttcgggccct cgggccggtg
     3421 cttgtggact cggctgcgac tggccaggga gatgtggatg tacagcaccg tcatgatgac
     3481 cacaggcagg tagaagccag caatggctgt gccaaaggtc actgctgggt tggacaggaa
     3541 ctggatgaag cactggttgt cgggcaccgt ccgcttaccc accacaaact gccagaacaa
     3601 gatggcaggc gcccagagca cgaaggacag tacccaggca gcagcaatca tgaggcctgc
     3661 catcttggtg gtgcgccggg cagggtaggt gagaggcttg gtgacgcaga agtagcggtc
     3721 aaagctgatg atgagaaggt tcatgacgga ggcgttgctc accacgcagt ccagggccag
     3781 ccacaggtcg cagaccacgg cgcccagggg ccagtagccc ttgatgatgt acacggtgta
     3841 gaggttcatg gagaaggcgc ctatgatgag atcagcacac gccaggctga agaggaagta
     3901 gttgttgact gtctgcagct gcctgttgac cttgatggac agcatcacca ggatgttgcc
     3961 cacgacagtc accaggctca gggagcctgt cactgtggca atgaagacca tttccaccgt
     4021 ctcatagcga ttgtgggatg atgacgtgac caggcgcacg gactgattgc ccgagctgcc
     4081 attgacaggt gtgaagttgg ccatggtggc gctagcataa cttcgtataa agtatcctat
     4141 acgaagttat ttgccttaac ccagaaatta tcactgttat tctttagaat ggtgcaaaga
     4201 ataacttcgt ataatgtatg ctatacgaag ttatgaattc cgctcgagat aatcaacctc
     4261 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     4321 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     4381 ttttctcctc cttgtataaa tcctggttag ttcttgccac ggcggaactc atcgccgcct
     4441 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttta
     4501 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat ctagctttat ttgtgaaatt
     4561 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
     4621 aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagcggc
     4681 cgcaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc tcgctcactg
     4741 aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg
     4801 agcgagcgcg cagctgcctg caggggcgcc tgatgcggta ttttctcctt acgcatctgt
     4861 gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta gcggcgcatt
     4921 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     4981 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     5041 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     5101 caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     5161 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     5221 aacactcaac tctatctcgg gctattcttt tgatttataa gggattttgc cgatttcggt
     5281 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     5341 aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     5401 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     5461 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     5521 gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa
     5581 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
     5641 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     5701 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     5761 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     5821 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     5881 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     5941 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     6001 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     6061 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     6121 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     6181 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     6241 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     6301 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     6361 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     6421 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     6481 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     6541 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     6601 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     6661 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     6721 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     6781 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     6841 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     6901 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     6961 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     7021 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     7081 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     7141 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     7201 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     7261 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     7321 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     7381 tttacggttc ctggcctttt gctggccttt tgctcacatg tt
//
