LOCUS       sequence-361677-        7307 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Cre-dependent AAV virus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7307)
  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 7307)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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: AiP5004 pAAV-Syn-DIO-TVA-dTom-SAD-B19G
FEATURES             Location/Qualifiers
     source          1..7307
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   33..162
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        190..637
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      644..653
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    659..692
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    743..776
                     /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(2357..2413)
                     /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(2423..3121)
                     /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(2506..2529)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(2704..2723)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2973..2991
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(3122..3178)
                     /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(3661..3694)
                     /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(3745..3778)
                     /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    3780..4368
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3833..3853)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4251..4262)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4376..4600
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4602..4742
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4602..4731
                     /label=AAV2 ITR
     rep_origin      4817..5272
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4904..4923)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5114..5135
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5289..5308)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5408..5430
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5468..5486)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5554..5658
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5659..5727,5728..6519)
                     /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(5877..5896)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6690..7278
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7179..7198
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctggccttt tgctggcctt ttgctcacat gtcctgcagg cagctgcgcg ctcgctcgct
       61 cactgaggcc gcccgggcgt cgggcgacct ttggtcgccc ggcctcagtg agcgagcgag
      121 cgcgcagaga gggagtggcc aactccatca ctaggggttc ctgctagact gcagagggcc
      181 ctgcgtatga gtgcaagtgg gttttaggac caggatgagg cggggtgggg gtgcctacct
      241 gacgaccgac cccgacccac tggacaagca cccaaccccc attccccaaa ttgcgcatcc
      301 cctatcagag agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc
      361 agcaccgcgg acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac
      421 tgaaggcgcg ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg gccaccttgg
      481 tcgcgtccgc gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg
      541 gcacgggcgc gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt
      601 gggcagcgga ggagtcgtgt cgtgcctgag agcgcagtca gccgccacca tggttaatat
      661 aacttcgtat aatgtatgct atacgaagtt atgcagaatg gtagctggat tgtagctgct
      721 attagcaata tgaaacctct taataacttc gtataaagta tcctatacga agttatggta
      781 cctacagtct ggtctcaccc ccactcttgt gtgattccca tgaagatatg atcttcccgc
      841 tttggggagt gactgacacc tccctccctg tccctctgag attgtgttgc gtaggttctg
      901 atcgattgac tcttctacaa catgtcatca ggaaaattat caacatcaag gcagtcaggg
      961 cccctgcact cagtaataca tacttccccc agttcgggag acccaagtca actcctgaga
     1021 cctgattgtg cacatcggga aggtgaactt caacaaaatc ctcagcctcg tcaccgtcct
     1081 tgaaaacggt agacgggtct gccagggggt gcacaagggg gataaccgag gattccaaca
     1141 actccatatg ttgctggagg agggatgatt gcatctctgg gattaagaca ttgccgtcag
     1201 gtcctaatat tataccattg aaaaacaccc cgttcacatg aggatgacac ctccccccaa
     1261 ctcttaaaca cccttttgaa gggaggatct cattccaagt tctgactgac ttgtagtgag
     1321 catcggcttc catcaaggtc ttgttgaata tggtatatgc ttttccaaac ccagggacaa
     1381 gttttcttaa atgactgaga cgtctgaaac tcactgactt ggttgtcatg atggactcta
     1441 gtgcatccag acactcctct ctcttcctga ccaactcctc tacaacaagg tgctcaattt
     1501 cgtctgagcg aaagtcgtgc aggttcacca acttatcggg agggcaccat ttggtttcat
     1561 ttgatgtttg catcgagacc catgttccat ccataagtct aagtcctaga actccacata
     1621 acttgagttt gcatgctcct tttaaagact tatataggcc tctttcatct acaaagccgc
     1681 aagtctcact ccctttggat gctctcttcc ctctactatt ggtaaaaatg tcacaagaca
     1741 tccctagtct cggattctcg ggcatccaaa tggtgtaatc gtggttagtg gagcagtagg
     1801 tagaagacac cgctactcct gagcacttcc cgctagggaa gaccctcgag tgaagggatc
     1861 tgtcatatgg gtccaaatct gccacacttg gagatatgat aacgagagac tccttggtgg
     1921 tttttacagt tcgaagccag cggtagtcag ggtacggatt gtgtagagac tcttcatatc
     1981 tggggtcacc ggccatcttc cagttgtacg cggctctaca tgcatctggt gttgggcgga
     2041 aatgctttct tttgaacgtg gttgtgacat aaccaacgaa gttagtgtag gtttcagcct
     2101 ccgtcacaac gcctgtgcaa gtgaacccgt tcacttttat ggctaagatg tatccaactt
     2161 taagttccat gtaggagaac cctgacaggt tggtgcatcc ttcgtcctcc actaccaaat
     2221 tgtttgggca gctgaggtga tgtatgtcaa tcggactcca gggaccaagc ttgtctggta
     2281 tcgtgtaaat agggaatttc ccaaaacaca atggaaaaac cagaaggggt acaaacagga
     2341 gagcctgagg aaccatgggt ccagggttct cctccacgtc tccagcctgc ttcagcaggc
     2401 tgaagttagt agctccggat cccttgtaca gctcgtccat gccgtacagg aacaggtggt
     2461 ggcggccctc ggagcgctcg tactgttcca cgatggtgta gtcctcgttg tgggaggtga
     2521 tgtccagctt ggtgtccacg tagtagtagc cgggcagttg cacgggcttc ttggccatgt
     2581 agatggtctt gaactccacc aggtagtggc cgccgtcctt cagcttcagg gcctggtgga
     2641 tctcgccctt cagcacgccg tcgcgggggt acaggcgctc ggtggaggcc tcccagccca
     2701 tggtcttctt ctgcattacg gggccgtcgg gggggaagtt ggtgccgcgc atcttcacct
     2761 tgtagatcag cgtgccgtcc tgcagggagg agtcctgggt cacggtcacc agaccgccgt
     2821 cctcgaagtt catcacgcgc tcccacttga agccctcggg gaaggacagc ttcttgtaat
     2881 cggggatgtc ggcggggtgc ttcacgtacg ccttggagcc gtacatgaac tggggggaca
     2941 ggatgtccca ggcgaagggc agggggccgc ccttggtcac cttcagcttg gcggtctggg
     3001 tgccctcgta ggggcggccc tcgccctcgc cctcgatctc gaactcgtgg ccgttcatgg
     3061 agccctccat gcgcaccttg aagcgcatga actctttgat gacctcctcg cccttgctca
     3121 cgggtccagg gttctcctcc acgtctccag cctgcttcag caggctgaag ttagtagctc
     3181 cggatccgga gaacaagtct gcctggctct tgtcaggcac cagcagctcc ttggatgcgc
     3241 tttcaagact gaagatgtca gacctgcttt ttgctttgga cttcccccat gcagcgatac
     3301 cacccacagc taccaggcag cacaggagca ctgcagtgat cagcatccac atgcggccgt
     3361 gattcctggc tggcagagca cgtccaggag caggggcagt gggagcctct gtgccgttgt
     3421 ccgtgggcac cgcggggatc gcgctggtcc cgcagcccca ctcgtcccgc ccgtcgtcgc
     3481 agtcggggtg tccgtcgcac agccagtcct gcgggtaaca ctccccgtgg gcaccgggcg
     3541 gctccgagca gcggaactga ccgggggggc aacgggacaa agaaccgtta ccggacccgt
     3601 taccggtcac gttaccgggc agcagcagca gcagcagcgc gggcagcagc cgcgcaccta
     3661 ataacttcgt atagcataca ttatacgaag ttatttgcct taacccagaa attatcactg
     3721 ttattcttta gaatggtgca aagaataact tcgtatagga tactttatac gaagttatga
     3781 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     3841 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     3901 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     3961 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     4021 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     4081 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     4141 tgggcactga caattccgtg gtgttgtcgg ggaagctgac gtcctttcca tggctgctcg
     4201 cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     4261 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     4321 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcct ggatcctgtg
     4381 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     4441 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     4501 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     4561 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg caggaacccc tagtgatgga
     4621 gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc
     4681 ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca
     4741 ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     4801 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     4861 acgcgcagcg tgaccgctac acttgccagc gccttagcgc ccgctccttt cgctttcttc
     4921 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     4981 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat
     5041 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     5101 acgttcttta atagtggact cttgttccaa actggaacaa cactcaactc tatctcgggc
     5161 tattcttttg atttataagg gattttgccg atttcggtct attggttaaa aaatgagctg
     5221 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc
     5281 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     5341 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     5401 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     5461 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     5521 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     5581 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     5641 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     5701 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     5761 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     5821 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     5881 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     5941 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     6001 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     6061 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     6121 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     6181 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     6241 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     6301 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     6361 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     6421 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     6481 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     6541 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     6601 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     6661 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     6721 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     6781 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     6841 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     6901 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     6961 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     7021 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     7081 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     7141 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     7201 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     7261 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggtt
//
