LOCUS       sequence-338888-        7100 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting envA-pseodotyped RVdG-CVS-N2c vectors for specific 
            retrograde labeling from cre-expressing inhibitory neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7100)
  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 7100)
  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-mDLX-TVA-2A-N2cG
FEATURES             Location/Qualifiers
     source          1..7100
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     intron          849..981
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     964..983
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     protein_bind    complement(1035..1068)
                     /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(1119..1152)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2752..2808)
                     /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(3265..3288)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      3288..3297
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3304..3337
                     /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    3388..3421
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3446..4034
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3499..3519)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3917..3928)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4203..4284
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4240..4259)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4294..4313
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   4363..4503
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4363..4492
                     /label=AAV2 ITR
     rep_origin      4578..5033
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4665..4684)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4875..4896
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5050..5069)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5169..5191
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5229..5247)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5315..5419
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5420..5488,5489..6280)
                     /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(5638..5657)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6451..7039
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6940..6959
                     /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 gcgtttaatt aagaattcta tacactcaca gtggtttggc
      181 atatatttgg tgaaattttt taaggaaaaa ttagtgttgg tttcgatata tggtagcttt
      241 ttctctaaca taatttgaat aattcagcaa agccctacta ccagctgtac ttctgcagcc
      301 tcttccattc tttccagcat tataattttg gttaattttc aattttaggt cctacgtctc
      361 tgcaatttgt gtatgaataa cagaataatt tccctctttt gtttcgcctt tcctgttcct
      421 gaatctaaat aaagatggct ttttagtatt aaaagtggaa gaaaattaca ggtaattatc
      481 tttgacggta aaaacgctgt aatcagcggg ctacatgaaa aattactcta attatggctg
      541 catttaagag aatggaaaaa aaccttcttg tggataaaaa ccttaaattg tccccaatgt
      601 ctgcttcaaa ttggatggca ctgcagctgg aggctttgtt cagaattgat cctggggagc
      661 tacgaaccca aagtttcaca gtaggaagct tgatatcgaa ttcctgcagc ccgggctggg
      721 cataaaagtc agggcagagc catctattgc ttacatttgc ttctagcctg caggtcgagg
      781 agcgcagcct tccagaagca gagcgcggcg ccttaagctg cagaagttgg tcgtgaggca
      841 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
      901 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
      961 actttgcctt tctctccaca ggtgtccact cccagttcaa ttacagctct taagaaacta
     1021 gagtcgactc cggaataact tcgtatagga tactttatac gaagttatgc agaatggtag
     1081 ctggattgta gctgctatta gcaatatgaa acctcttaat aacttcgtat agcatacatt
     1141 atacgaagtt atggcgcgcc ttacttgtac atctcacagt ctgatctcac ctccactctt
     1201 atatgattcc catgaaggta tgacttttcc gctttgggaa gtgactgaca cattcccccc
     1261 tgtccctcca aaactgcgtt gtttcgattc tggtcgattg gctcttctgc accatgtcat
     1321 tagggaaaat atcaacacca ggccaatcat ggcccctgca gtcatcaata catactttcc
     1381 ccagttcggg agacccaggt caacccctga gatctgtttg tacacatcgg ggaggtgaac
     1441 ttcaacaaaa tcctcagcct catcaccttc tttgaaaact gtagaagggt ctgccagggg
     1501 gtgcatcagg gggataactg aagattccaa caactccata tgttgctgga ggagggatga
     1561 ttgcatctct gggattagga catggtcgtc aggccctaat attataccat tgaaaaacac
     1621 cccgttcaca tgaggatggc accttcctcc aactttcaaa cacccttttg aggggatgat
     1681 ctcattccag gtccggactg acttgtagtg agcatcagcc tccatcaagg ttttgttgaa
     1741 tatggtatat gcttttccaa accctgggac aagttttctc aggtgactga gacgtctgaa
     1801 acttactgac ttggtggtca tgatggactc taatgcgtcc agacattcct ctcttttctt
     1861 gactaactcc tccacaacga gatgctcaat ctcgtctgag cgaaagtcgt gcaaattcac
     1921 caactgatct ggagggcacc atttggtctc atctgatgtt tgcatcgcga cccatgttcc
     1981 atccataagt ctaagtccaa gaactccaca taacttgagc ctgcatgctc cttttagaga
     2041 cttatacagg cctctttcat ccacaaagcc gcaagtcttg ttcccgttgg atgctctctt
     2101 ccctctgcta ttggtaaaaa tgtcacaagg tgtccttggt ctcggattct cgggcatcca
     2161 aatggtgtaa tcatggttag ttgagcagta ggtagaggac accgttattc ctgagcactt
     2221 tccgccaggg aagacccttg agtgaaggga tttgtcatat gggtccaaat ctgtcacact
     2281 tggggatatg ataatgaggg actctttggt ggttcttaca gttcgaagcc agtggtagtc
     2341 ggggtatgga ttgtgtaggg actcttcata tctggggtca ccggccatct tccagttata
     2401 cgcggctcta catgcgtctg gggtggggcg gaaatgcttt ctcttgaatg tggttgtgac
     2461 ataaccaaca aagttggtgt aggtctctgc ctctgtcaca acacctgtgc aagtgaaccc
     2521 gttcactttg atggctgaga tgtatcccac tttgagttcc atgtaggaga actcggacag
     2581 gttggtacat ccttcatcct ccacaaccag gttatttgga cagctgagat ggtgtatgtc
     2641 aatagggctc cagggaccaa gttcgtctgg tatcgtgtaa atggggaact tcccgaaaca
     2701 caacgaaaaa cccagaagga gtacaaacaa aagaacctga ggaaccatgg ttggcccggg
     2761 attctcttcg acatcccctg cttgtttcaa cagggagaag ttagtggctc cgcttccgtt
     2821 gtcttcgaac ccacccccct tggatgcgct ttcaagactg aagatgtcag acctgctttt
     2881 tgctttggac ttcccccatg cagcgatacc acccacagct accaggcagc acaggagcac
     2941 tgcagtgatc agcatccaca tgcggccgtg attcctggct ggcagagcac gtccaggagc
     3001 aggggcagtg ggagcctctg tgccgttgtc cgtgggcacc gcggggatcg cgctggtccc
     3061 gcagccccac tcgtcccgcc cgtcgtcgca gtcggggtgt ccgtcgcaca gccagtcctg
     3121 cgggtaacac tccccgtggg caccgggcgg ctccgagcag cggaactgac cgggggggca
     3181 acgggacaaa gaaccgttac cggacccgtt accggtcacg ttaccgggca gcagcagcag
     3241 cagcagcgcg ggcagcagcc gcgccttgtc gtcatcgtct ttgtagtcca tggtggcact
     3301 agcataactt cgtataaagt atcctatacg aagttatttg ccttaaccca gaaattatca
     3361 ctgttattct ttagaatggt gcaaagaata acttcgtata atgtatgcta tacgaagtta
     3421 tgaattcgat atcaagctta tcgataatca acctctggat tacaaaattt gtgaaagatt
     3481 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     3541 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     3601 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     3661 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     3721 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     3781 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
     3841 atcatcgtcc tttccttggc tgctcgccta tgttgccacc tggattctgc gcgggacgtc
     3901 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
     3961 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
     4021 ggccgcctcc ccgcatcgat accgagcgct gctcgagaga tctgtgatag cggccatcaa
     4081 gctggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
     4141 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
     4201 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4261 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4321 cagcttatcg ataccgcatg cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg
     4381 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     4441 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc
     4501 aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     4561 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     4621 tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt
     4681 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     4741 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga
     4801 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     4861 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg
     4921 ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct
     4981 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg
     5041 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     5101 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     5161 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     5221 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     5281 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     5341 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     5401 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     5461 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     5521 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     5581 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     5641 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     5701 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     5761 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     5821 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     5881 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5941 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     6001 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     6061 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     6121 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     6181 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     6241 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     6301 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     6361 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     6421 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     6481 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     6541 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     6601 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     6661 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     6721 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     6781 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     6841 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     6901 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     6961 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     7021 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     7081 ctggcctttt gctcacatgt
//
