LOCUS       sequence-338741-        7687 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting envA-pseodotyped RVdG-CVS-N2c vectors for specific 
            retrograde labeling specifically from cre-expressing neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7687)
  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 7687)
  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-DIO-Ef1a-TVA-2A-oG
FEATURES             Location/Qualifiers
     source          1..7687
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1440..1473)
                     /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)."
     protein_bind    complement(1524..1557)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(3144..3200)
                     /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(3633..3656)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      3656..3665
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3672..3705
                     /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)."
     protein_bind    3756..3789
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3814..4402
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3867..3887)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4285..4296)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4434..4910
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4633..4652)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4950..5090
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4950..5079
                     /label=AAV2 ITR
     rep_origin      5165..5620
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5252..5271)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5462..5483
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5637..5656)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5756..5778
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5816..5834)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5902..6006
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6007..6075,6076..6867)
                     /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(6225..6244)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7038..7626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7527..7546
                     /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 gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggaa
     1441 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
     1561 gcgccttaga gccgtgtctc gcccccagat ttatgacttt cccaactgct gatgatttta
     1621 ccgctctggg gtgtgacaga cacttctcta ccggtccccc tcagattgtg ctgagttggc
     1681 tcggagcggt tcactcgtcg ccagcaggtc atcaggaaaa tgatcagcat gagagcagtc
     1741 agtgctccag cggagagcag cacgtacttg ccccaattag gcagtccgag atcgacgcca
     1801 ctgatccgtt cgtgcacgtc gggcaggtgg acctccacaa aatcttcggc ctcgtcgccg
     1861 tttttgaaca ctgtgcttgg gtctgcgaga ggatgcatca ggggaatgac gcttgacacg
     1921 agcagttcca tgtgctgctg gagcagacta gactgcatct cggggatcag cacgttacca
     1981 tctggcccca gaatgatacc attaaagaag accccgttca catgtgggtg acatcgaccc
     2041 ccgactcgca ggcatccctt actaggaatg atttcattcc aggttctcac agatttataa
     2101 tgggcgtctg cctccatcag tgtcttgtta aagatggtgt atgcttttcc gaagccaggg
     2161 acgagcttcc tcaggtggct cagcctcctg aagctcacgc tcttggtagt cataattgac
     2221 tccagagcat cgagacattc ctcccttttc ttcacgagtt cctcgaccac cagatgttcg
     2281 atctcatcgc ttcggaagtc gtgcagattc acgagctgcc caggggggca ccacttagtc
     2341 tcgttagatg tctgcatagc gacccaggtt ccgtccatca gccggagtcc cagcacgcca
     2401 cagagtttca gcttgcaggc acctttgagg ctcttataca gccctctttc atccacaaat
     2461 ccacaagtct cgctgccttt tgaggccctc ttgcccctgc tgttggtgaa gatgtcgcag
     2521 gacatcccca gccttggatt ctcaggcatc cagattgtat aatcatggtt ggtggaacag
     2581 taagtggagc tgacggccac tccggagcag tttccgccgg gaaacactgg actgtggaga
     2641 gatcggtcgt aggggtccag gtcggcgaca gatggggaaa tgatgaccag gctctctttt
     2701 gtggtcttca cagtgcgcag ccaatgatag tctgggtaag gattgtgcag gctttcctca
     2761 tagcgggggt cgccagccat cttccagttg taagctgccc gacaggcgtc tggtgttggc
     2821 cgaaagtgtt ttctcttgaa agttgtggtg acatatccca caaaatttgt gtaggtttct
     2881 gcctcggtga ccacaccagt gcatgtgaac ccgttcatct taatagcaga gatatagccc
     2941 actttcagct ccatgtagga gaatccgctc aggttggtac agccttcgtc ctcgaccacg
     3001 agattgttag ggcatgacag atggtgaata tcgataggac tccaggggcc cagcttgtcg
     3061 gggatagtgt agatagggaa tttaccaaag cagagtggga agacgagcag agggacgaag
     3121 aggagagcct gtgggaccat ggttggcccg ggattctctt cgacatcccc tgcttgtttc
     3181 aacagggaga agttagtggc tccgcttccg ttgtcttcga acccaccccc cttggatgcg
     3241 ctttcaagac tgaagatgtc agacctgctt tttgctttgg acttccccca tgcagcgata
     3301 ccacccacag ctaccaggca gcacaggagc actgcagtga tcagcatcca catgcggccg
     3361 tgattcctgg ctggcagagc acgtccagga gcaggggcag tgggagcctc tgtgccgttg
     3421 tccgtgggca ccgcggggat cgcgctggtc ccgcagcccc actcgtcccg cccgtcgtcg
     3481 cagtcggggt gtccgtcgca cagccagtcc tgcgggtaac actccccgtg ggcaccgggc
     3541 ggctccgagc agcggaactg accggggggg caacgggaca aagaaccgtt accggacccg
     3601 ttaccggtca cgttaccggg cagcagccgc gccttgtcgt catcgtcttt gtagtccatg
     3661 gtggcactag cataacttcg tataaagtat cctatacgaa gttatttgcc ttaacccaga
     3721 aattatcact gttattcttt agaatggtgc aaagaataac ttcgtataat gtatgctata
     3781 cgaagttatg aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt
     3841 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     3901 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     3961 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     4021 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     4081 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     4141 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     4201 tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg gattctgcgc
     4261 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     4321 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     4381 tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc tacgggtggc
     4441 atccctgtga cccctcccca gtgcctctcc tggccctgga agttgccact ccagtgccca
     4501 ccagccttgt cctaataaaa ttaagttgca tcattttgtc tgactaggtg tccttctata
     4561 atattatggg gtggaggggg gtggtatgga gcaaggggca agttgggaag acaacctgta
     4621 gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt ggcacaatct tggctcactg
     4681 caatctccgc ctcctgggtt caagcgattc tcctgcctca gcctcccgag ttgttgggat
     4741 tccaggcatg catgaccagg ctcagctaat ttttgttttt ttggtagaga cggggtttca
     4801 ccatattggc caggctggtc tccaactcct aatctcaggt gatctaccca ccttggcctc
     4861 ccaaattgct gggattacag gcgtgaacca ctgctccctt ccctgtcctt ctgattttgt
     4921 aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc
     4981 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     5041 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat
     5101 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
     5161 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     5221 accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc
     5281 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     5341 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
     5401 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     5461 agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat
     5521 ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa
     5581 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
     5641 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     5701 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     5761 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     5821 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     5881 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     5941 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     6001 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     6061 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     6121 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     6181 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     6241 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     6301 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     6361 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     6421 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     6481 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     6541 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     6601 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     6661 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6721 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6781 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6841 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     6901 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     6961 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     7021 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     7081 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     7141 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     7201 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     7261 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     7321 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     7381 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     7441 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     7501 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     7561 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     7621 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     7681 cacatgt
//
