LOCUS       sequence-350219-        8094 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting envA-pseodotyped RVdG-CVS-N2c vectors for specific 
            retrograde labeling from excitatory cortical neurons which do not 
            express cre-recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8094)
  AUTHORS   Sumser A, Joesch M, Jonas P, Ben-Simon Y
  TITLE     Fast, high-throughput production of improved rabies viral vectors 
            for specific, efficient and versatile transsynaptic retrograde 
            labeling.
  JOURNAL   Elife. 2022 Aug 30;11:e79848. doi: 10.7554/eLife.79848.
  PUBMED    36040301
REFERENCE   2  (bases 1 to 8094)
  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"; journalName: "Elife."; 
            date: "2022-08-30"; volume: "11:e79848. doi"; pages: " 
            10.7554/eLife.79848"
COMMENT     Sequence Label: pAAV-SEO-CaMKII-TVA-P2A-N2cG
FEATURES             Location/Qualifiers
     source          1..8094
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        179..197
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     179..196
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     repeat_region   218..347
                     /label=AAV2 ITR
     protein_bind    complement(428..461)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        500..1788
                     /label=CaMKII promoter
                     /note="promoter for the mouse calcium/calmodulin-dependent 
                     kinase II alpha subunit"
     regulatory      1815..1824
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2280..2345
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             2367..2423
                     /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(4042..4075)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4131..4719
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4184..4204)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4602..4613)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4746..4763)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4752..4976
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(5186..5515)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5200..5335
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     rep_origin      complement(5761..6349)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5841..5860)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6520..7311,7312..7380))
                     /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     7143..7162
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7381..7485)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7553..7571
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7609..7631)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7731..7750
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7964..7983
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 ggacggcgtt accagaaact cagaaggttc gtccaaccaa accgactctg acggcagttt
       61 acgagagaga tgatagggtc tgcttcagta agccagatgc tacacaatta ggcttgtaca
      121 tattgtcgtt agaacgcggc tacaattaat acataacctt atgtatcata cacatacgat
      181 ttaggtgaca ctatagaata cacggaatta attctagctg cgcgctcgct cgctcactga
      241 ggccgcccgg gcaaagcccg ggcgtcgggc gacctttggt cgcccggcct cagtgagcga
      301 gcgagcgcgc agagagggag tggccaactc catcactagg ggttccttgt agttaatgat
      361 taacccgcca tgctacttat ctacgtagcc atgctctagg taccattcca gcaggtcgag
      421 ggacctaata acttcgtata gcatacatta tacgaagtta tattaagggt tccaagctta
      481 agcggccgca cgcgtttaac attatggcct taggtcactt catctccatg gggttcttct
      541 tctgattttc tagaaaatga gatgggggtg cagagagctt cctcagtgac ctgcccaggg
      601 tcacatcaga aatgtcagag ctagaacttg aactcagatt actaatctta aattccatgc
      661 cttgggggca tgcaagtacg atatacagaa ggagtgaact cattagggca gatgaccaat
      721 gagtttagga aagaagagtc cagggcaggg tacatctaca ccacccgccc agccctgggt
      781 gagtccagcc acgttcacct cattatagtt gcctctctcc agtcctacct tgacgggaag
      841 cacaagcaga aactgggaca ggagccccag gagaccaaat cttcatggtc cccctgggag
      901 gatgggtggg gagagctgtg gcagaggcct caggaggggc cctgctgctc agtggtgaca
      961 gataggggtg agaaagcaga cagagtcatt ccgtcagcat tctgggtctg tttggtactt
     1021 cttctcacgc taaggtggcg gtgtgatatg cacaatggct aaaaagcagg gagagctgga
     1081 aagaaacaag gacagagaca gaggccaagt caaccagacc aattcccaga ggaagcaaag
     1141 aaaccattac agagactaca agggggaagg gaaggagaga tgaattagct tcccctgtaa
     1201 accttagaac ccagctgttg ccagggcaac ggggcaatac ctgtctcttc agaggagatg
     1261 aagttgccag ggtaactaca tcctgtcttt ctcaaggacc atcccagaat gtggcaccca
     1321 ctagccgtta ccatagcaac tgcctctttg ccccacttaa tcccatcccg tctgttaaaa
     1381 gggccctata gttggaggtg ggggaggtag gaagagcgat gatcacttgt ggactaagtt
     1441 tgttcgcatc cccttctcca accccctcag tacatcaccc tgggggaaca gggtccactt
     1501 gctcctgggc ccacacagtc ctgcagtatt gtgtatataa ggccagggca aagaggagca
     1561 ggttttaaag tgaaaggcag gcaggtgttg gggaggcagt taccggggca acgggaacag
     1621 ggcgtttcgg aggtggttgc catggggacc tggatgctga cgaaggctcg cgaggctgtg
     1681 agcagccaca gtgccctgct cagaagcccc aagctcgtca gtcaagccgg ttctccgttt
     1741 gcactcagga gcacgggcag gcgagtggcc cctagttctg ggggcagctc tagagcggta
     1801 ccggatccac tagtgccacc atggcgcggc tgctgcccgc gctgctgctg ctgctgctgc
     1861 ccggtaacgt gaccggtaac gggtccggta acggttcttt gtcccgttgc ccccccggtc
     1921 agttccgctg ctcggagccg cccggtgccc acggggagtg ttacccgcag gactggctgt
     1981 gcgacggaca ccccgactgc gacgacgggc gggacgagtg gggctgcggg accagcgcga
     2041 tccccgcggt gcccacggac aacggcacag aggctcccac tgcccctgct cctggacgtg
     2101 ctctgccagc caggaatcac ggccgcatgt ggatgctgat cactgcagtg ctcctgtgct
     2161 gcctggtagc tgtgggtggt atcgctgcat gggggaagtc caaagcaaaa agcaggtctg
     2221 acatcttcag tcttgaaagc gcatccaagg ggggtgggtt cgaaccgggc agcaccggcg
     2281 attataagga tcatgatggt gattataagg atcatgatat cgactacaaa gacgatgacg
     2341 acaagttcga agacaacgga agcggagcca ctaacttctc cctgttgaaa caagcagggg
     2401 atgtcgaaga gaatcccggg ccaaccatgg ttcctcaggt tcttttgttt gtactccttc
     2461 tgggtttttc gttgtgtttc gggaagttcc ccatttacac gataccagac gaacttggtc
     2521 cctggagccc tattgacata caccatctca gctgtccaaa taacctggtt gtggaggatg
     2581 aaggatgtac caacctgtcc gagttctcct acatggaact caaagtggga tacatctcag
     2641 ccatcaaagt gaacgggttc acttgcacag gtgttgtgac agaggcagag acctacacca
     2701 actttgttgg ttatgtcaca accacattca agagaaagca tttccgcccc accccagacg
     2761 catgtagagc cgcgtataac tggaagatgg ccggtgaccc cagatatgaa gagtccctac
     2821 acaatccata ccccgactac cactggcttc gaactgtaag aaccaccaaa gagtccctca
     2881 ttatcatatc cccaagtgtg acagatttgg acccatatga caaatccctt cactcaaggg
     2941 tcttccctgg cggaaagtgc tcaggaataa cggtgtcctc tacctactgc tcaactaacc
     3001 atgattacac catttggatg cccgagaatc cgagaccaag gacaccttgt gacattttta
     3061 ccaatagcag agggaagaga gcatccaacg ggaacaagac ttgcggcttt gtggatgaaa
     3121 gaggcctgta taagtctcta aaaggagcat gcaggctcaa gttatgtgga gttcttggac
     3181 ttagacttat ggatggaaca tgggtcgcga tgcaaacatc agatgagacc aaatggtgcc
     3241 ctccagatca gttggtgaat ttgcacgact ttcgctcaga cgagattgag catctcgttg
     3301 tggaggagtt agtcaagaaa agagaggaat gtctggacgc attagagtcc atcatgacca
     3361 ccaagtcagt aagtttcaga cgtctcagtc acctgagaaa acttgtccca gggtttggaa
     3421 aagcatatac catattcaac aaaaccttga tggaggctga tgctcactac aagtcagtcc
     3481 ggacctggaa tgagatcatc ccctcaaaag ggtgtttgaa agttggagga aggtgccatc
     3541 ctcatgtgaa cggggtgttt ttcaatggta taatattagg gcctgacgac catgtcctaa
     3601 tcccagagat gcaatcatcc ctcctccagc aacatatgga gttgttggaa tcttcagtta
     3661 tccccctgat gcaccccctg gcagaccctt ctacagtttt caaagaaggt gatgaggctg
     3721 aggattttgt tgaagttcac ctccccgatg tgtacaaaca gatctcaggg gttgacctgg
     3781 gtctcccgaa ctggggaaag tatgtattga tgactgcagg ggccatgatt ggcctggtgt
     3841 tgatattttc cctaatgaca tggtgcagaa gagccaatcg accagaatcg aaacaacgca
     3901 gttttggagg gacagggggg aatgtgtcag tcacttccca aagcggaaaa gtcatacctt
     3961 catgggaatc atataagagt ggaggtgaga tcagactgta agatgtacaa gtaaggcgcg
     4021 ccgaattcgt cgagggacct aataacttcg tatagcatac attatacgaa gttatacatg
     4081 tttaagggtt ccggttccac taggtacaat tcgatatcaa gcttatcgat aatcaacctc
     4141 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     4201 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     4261 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
     4321 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
     4381 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
     4441 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
     4501 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
     4561 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
     4621 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
     4681 ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccgt cgactcgctg
     4741 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     4801 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     4861 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     4921 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     4981 tgaggcggaa agaaccagct ggggctcgac tagagcatgg ctacgtagat aagtagcatg
     5041 gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc
     5101 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggcggcctca
     5161 gtgagcgagc gagcgcgcag agctttttgc aaaagcctag gcctccaaaa aagcctcctc
     5221 actacttctg gaatagctca gaggccgagg cggcctcggc ctctgcataa ataaaaaaaa
     5281 ttagtcagcc atggggcgga gaatgggcgg aactgggcgg agttaggggc gggatgggcg
     5341 gagttagggg cgggactatg gttgctgact aattgagatg catgctttgc atacttctgc
     5401 ctgctgggga gcctggggac tttccacacc tggttgctga ctaattgaga tgcatgcttt
     5461 gcatacttct gcctgctggg gagcctgggg actttccaca ccctaactga cacacgttcc
     5521 acagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     5581 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     5641 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5701 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5761 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5821 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5881 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5941 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     6001 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     6061 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     6121 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     6181 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     6241 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     6301 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     6361 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     6421 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     6481 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     6541 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     6601 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6661 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6721 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6781 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6841 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6901 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6961 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7021 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7081 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     7141 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7201 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     7261 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     7321 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     7381 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     7441 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     7501 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     7561 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     7621 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     7681 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     7741 gcagattgta ctgagagtgc accattcgac gctctccctt atgcgactcc tgcattagga
     7801 agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca
     7861 aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac
     7921 aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat
     7981 aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga
     8041 ggatctggct agcgatgacc ctgctgattg gttcgctgac catttccggg tgcg
//
