LOCUS       sequence-361679-        7308 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent AAV virus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7308)
  AUTHORS   Yao S, Wang Q, Hirokawa KE, Ouellette B, Ahmed R, Bomben J, Brouner 
            K, Casal L, Caldejon S, Cho A, Dotson NI, Daigle TL, Egdorf T, 
            Enstrom R, Gary A, Gelfand E, Gorham M, Griffin F, Gu H, Hancock N, 
            Howard R, Kuan L, Lambert S, Lee EK, Luviano J, Mace K, Maxwell M, 
            Mortrud MT, Naeemi M, Nayan C, Ngo NK, Nguyen T, North K, Ransford 
            S, Ruiz A, Seid S, Swapp J, Taormina MJ, Wakeman W, Zhou T, Nicovich
            PR, Williford A, Potekhina L, McGraw M, Ng L, Groblewski PA, Tasic 
            B, Mihalas S, Harris JA, Cetin A, Zeng H
  TITLE     A whole-brain monosynaptic input connectome to neuron classes in 
            mouse visual cortex.
  JOURNAL   Nat Neurosci. 2023 Feb;26(2):350-364. doi: 
            10.1038/s41593-022-01219-x. Epub 2022 Dec 22.
  PUBMED    36550293
REFERENCE   2  (bases 1 to 7308)
  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"; doi: 
            "10.1038/s41593-022-01219-x"; journalName: "Nat Neurosci"; date: 
            "2023-02"; volume: "26"; issue: "2"; pages: "350-364"
COMMENT     Sequence Label: AiP5003 pAAV-Syn-DIO-TVA66T-dTom-CVS-N2cG
FEATURES             Location/Qualifiers
     source          1..7308
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        59..506
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      513..522
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    528..561
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    612..645
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     CDS             complement(2227..2283)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(2293..2991)
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGL
                     VTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIHQ
                     ALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRHH
                     LFLYGMDELYK"
     primer_bind     complement(2376..2399)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(2574..2593)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2843..2861
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2992..3048)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     protein_bind    complement(3531..3564)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(3615..3648)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     misc_feature    3650..4238
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3703..3723)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4121..4132)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4246..4470
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4472..4612
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4472..4601
                     /label=AAV2 ITR
     rep_origin      4687..5142
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4774..4793)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4984..5005
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5159..5178)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5278..5300
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5338..5356)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5424..5528
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5529..5597,5598..6389)
                     /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(5747..5766)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6560..7148
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7049..7068
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   join(7210..7308,1..31)
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 ggagtggcca actccatcac taggggttcc tgctagactg cagagggccc tgcgtatgag
       61 tgcaagtggg ttttaggacc aggatgaggc ggggtggggg tgcctacctg acgaccgacc
      121 ccgacccact ggacaagcac ccaaccccca ttccccaaat tgcgcatccc ctatcagaga
      181 gggggagggg aaacaggatg cggcgaggcg cgtgcgcact gccagcttca gcaccgcgga
      241 cagtgccttc gcccccgcct ggcggcgcgc gccaccgccg cctcagcact gaaggcgcgc
      301 tgacgtcact cgccggtccc ccgcaaactc cccttcccgg ccaccttggt cgcgtccgcg
      361 ccgccgccgg cccagccgga ccgcaccacg cgaggcgcga gatagggggg cacgggcgcg
      421 accatctgcg ctgcggcgcc ggcgactcag cgctgcctca gtctgcggtg ggcagcggag
      481 gagtcgtgtc gtgcctgaga gcgcagtcag ccgccaccat ggttaatata acttcgtata
      541 atgtatgcta tacgaagtta tgcagaatgg tagctggatt gtagctgcta ttagcaatat
      601 gaaacctctt aataacttcg tataaagtat cctatacgaa gttatggtac ctcagagtct
      661 tatttcaccc ccggatttgt agctctccca gctgggtatc accttcccag attgagatgt
      721 aacactcacg ttacccccgg tccccccaaa ggacctctgc ttggattcgg gccggtttgc
      781 tctccgacac caagtcatga gactgaaaat aagtaccaaa ccaatcatag ctccggcagt
      841 catcaaaacg tatttgcccc agtttggcaa accgagatcc actccgctga tttgtttata
      901 gacatcaggc aggtgtacct ctacaaagtc ctcagcttca tcgccttctt tgaaaacagt
      961 gcttggatca gccaaaggat gcataagagg aatcacgctg ctttccaaca gctccatgtg
     1021 ttgttgaagg aggctagact gcatctcagg aatcaaaaca tggtcatcgg gtccgagaat
     1081 gatgccgtta aaaaaaacgc cgttcacgtg tgggtgacac cttccgccaa ctttaagaca
     1141 tcctttgcta ggtatgattt cgttccaagt gcgtacagat ttgtagtgag catcagcttc
     1201 cataagagtc ttgttgaata tggtataagc tttgccgaaa ccagggacca actttctcag
     1261 gtgagacagt ctccggaagc taacagactt tgttgtcatg attgattcca gtgcgtccag
     1321 gcattcttcc cgcttcttca ccaactcttc aaccacgaga tgttcaattt catcgcttcg
     1381 gaagtcatgc aaatttacga gctgatcggg ggggcaccat ttagtctcat cactggtttg
     1441 catagctacc caagttccat ccatgagcct gagtccaaga acaccgcaca gtttcagtct
     1501 acaggcaccc ttgagactct tatacaggcc gcgttcgtct acgaaaccac aggtcttgtt
     1561 gccgttagaa gctcgtttac cgcgagagtt tgtaaaaata tcgcaggggg tccggggtct
     1621 gggattttct ggcatccaga tagtgtagtc gtggtttgtt gaacagtaag tggatgaaac
     1681 agttattccg ctacatttgc ccccgggaaa gactctagaa tgaagacttt tgtcatatgg
     1741 gtccaaatcg gtcacggagg gactgatgat aatcaaactt tcttttgtgg tccttactgt
     1801 tcggagccaa tggtaatctg gatagggatt gtgcagtgac tcttcgtacc ttgggtcacc
     1861 ggccattttc caattatatg cagcccgaca agcatcagga gtaggtcgaa aatgtttgcg
     1921 cttaaaagtg gtagtcacgt atccgacgaa attagtgtag gtctcggcct cggttacgac
     1981 gccggtacat gtaaagccgt tgactttgat agcggaaata taaccaacct tgagttccat
     2041 gtatgaaaat tcgctgagat tggtacaacc ctcatcctcg acgaccaggt tgttgggaca
     2101 tgagaggtgg tggatgtcga taggagacca aggtccaagc tcatcgggaa tagtataaat
     2161 tggaaatttg ccgaaacaga gactaaatcc aaggagaagt acaaaaagga gtacctgagg
     2221 aaccatgggt ccagggttct cctccacgtc tccagcctgc ttcagcaggc tgaagttagt
     2281 agctccggat cccttgtaca gctcgtccat gccgtacagg aacaggtggt ggcggccctc
     2341 ggagcgctcg tactgttcca cgatggtgta gtcctcgttg tgggaggtga tgtccagctt
     2401 ggtgtccacg tagtagtagc cgggcagttg cacgggcttc ttggccatgt agatggtctt
     2461 gaactccacc aggtagtggc cgccgtcctt cagcttcagg gcctggtgga tctcgccctt
     2521 cagcacgccg tcgcgggggt acaggcgctc ggtggaggcc tcccagccca tggtcttctt
     2581 ctgcattacg gggccgtcgg gggggaagtt ggtgccgcgc atcttcacct tgtagatcag
     2641 cgtgccgtcc tgcagggagg agtcctgggt cacggtcacc agaccgccgt cctcgaagtt
     2701 catcacgcgc tcccacttga agccctcggg gaaggacagc ttcttgtaat cggggatgtc
     2761 ggcggggtgc ttcacgtacg ccttggagcc gtacatgaac tggggggaca ggatgtccca
     2821 ggcgaagggc agggggccgc ccttggtcac cttcagcttg gcggtctggg tgccctcgta
     2881 ggggcggccc tcgccctcgc cctcgatctc gaactcgtgg ccgttcatgg agccctccat
     2941 gcgcaccttg aagcgcatga actctttgat gacctcctcg cccttgctca cgggtccagg
     3001 gttctcctcc acgtctccag cctgcttcag caggctgaag ttagtagctc cggatccgga
     3061 gaacaagtct gcctggctct tgtcaggcac cagcagctcc ttggatgcgc tttcaagact
     3121 gaagatgtca gacctgcttt ttgctttgga cttcccccat gcagcgatac cacccacagc
     3181 taccaggcag cacaggagca ctgcagtgat cagcatccac atgcggccgt gattcctggc
     3241 tggcagagca cgtccaggag caggggcagt gggagcctct gtgccgttgt ccgtgggcac
     3301 cgcggggatc gcgctggtcc cgcagcccca ggtgtcccgc ccgtcgtcgc agtcggggtg
     3361 tccgtcgcac agccagtcct gcgggtaaca ctccccgtgg gcaccgggcg gctccgagca
     3421 gcggaactga ccgggggggc aacgggacaa agaaccgtta ccggacccgt taccggtcac
     3481 gttaccgggc agcagcagca gcagcagcgc gggcagcagc cgcgcaccta ataacttcgt
     3541 atagcataca ttatacgaag ttatttgcct taacccagaa attatcactg ttattcttta
     3601 gaatggtgca aagaataact tcgtatagga tactttatac gaagttatga atcaacctct
     3661 ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct
     3721 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat
     3781 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt
     3841 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat
     3901 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc
     3961 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
     4021 caattccgtg gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg cctgtgttgc
     4081 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga
     4141 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc
     4201 tcagacgagt cggatctccc tttgggccgc ctccccgcct ggatcctgtg ccttctagtt
     4261 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     4321 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     4381 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     4441 ggcatgctgg ggatgcggtg ggctctatgg caggaacccc tagtgatgga gttggccact
     4501 ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg
     4561 ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg
     4621 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac
     4681 catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     4741 tgaccgctac acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc
     4801 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     4861 gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta
     4921 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     4981 atagtggact cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg
     5041 atttataagg gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa
     5101 aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta
     5161 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     5221 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     5281 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     5341 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     5401 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     5461 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     5521 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     5581 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     5641 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     5701 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     5761 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     5821 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     5881 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     5941 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     6001 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     6061 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     6121 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     6181 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     6241 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     6301 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     6361 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     6421 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     6481 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     6541 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     6601 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     6661 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     6721 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     6781 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     6841 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     6901 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     6961 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     7021 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     7081 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     7141 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     7201 ctcacatgtc ctgcaggcag ctgcgcgctc gctcgctcac tgaggccgcc cgggcgtcgg
     7261 gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagag
//
