LOCUS       sequence-338887-        6920 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Targeting envA-pseodotyped RVdG-CVS-N2c vectors for specific 
            retrograde labeling from inhibitory neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6920)
  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 6920)
  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-mDLX-TVA-2A-N2cG
FEATURES             Location/Qualifiers
     source          1..6920
                     /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"
     regulatory      1023..1032
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1488..1553
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1575..1631
                     /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"
     misc_feature    3266..3854
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3319..3339)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3737..3748)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4023..4104
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4060..4079)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4114..4133
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   4183..4323
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4183..4312
                     /label=AAV2 ITR
     rep_origin      4398..4853
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4485..4504)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4695..4716
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4870..4889)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4989..5011
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5049..5067)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5135..5239
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5240..5308,5309..6100)
                     /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(5458..5477)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6271..6859
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6760..6779
                     /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 gtgccaccat ggcgcggctg ctgcccgcgc tgctgctgct gctgctgccc ggtaacgtga
     1081 ccggtaacgg gtccggtaac ggttctttgt cccgttgccc ccccggtcag ttccgctgct
     1141 cggagccgcc cggtgcccac ggggagtgtt acccgcagga ctggctgtgc gacggacacc
     1201 ccgactgcga cgacgggcgg gacgagtggg gctgcgggac cagcgcgatc cccgcggtgc
     1261 ccacggacaa cggcacagag gctcccactg cccctgctcc tggacgtgct ctgccagcca
     1321 ggaatcacgg ccgcatgtgg atgctgatca ctgcagtgct cctgtgctgc ctggtagctg
     1381 tgggtggtat cgctgcatgg gggaagtcca aagcaaaaag caggtctgac atcttcagtc
     1441 ttgaaagcgc atccaagggg ggtgggttcg aaccgggcag caccggcgat tataaggatc
     1501 atgatggtga ttataaggat catgatatcg actacaaaga cgatgacgac aagttcgaag
     1561 acaacggaag cggagccact aacttctccc tgttgaaaca agcaggggat gtcgaagaga
     1621 atcccgggcc aaccatggtt cctcaggttc ttttgtttgt actccttctg ggtttttcgt
     1681 tgtgtttcgg gaagttcccc atttacacga taccagacga acttggtccc tggagcccta
     1741 ttgacataca ccatctcagc tgtccaaata acctggttgt ggaggatgaa ggatgtacca
     1801 acctgtccga gttctcctac atggaactca aagtgggata catctcagcc atcaaagtga
     1861 acgggttcac ttgcacaggt gttgtgacag aggcagagac ctacaccaac tttgttggtt
     1921 atgtcacaac cacattcaag agaaagcatt tccgccccac cccagacgca tgtagagccg
     1981 cgtataactg gaagatggcc ggtgacccca gatatgaaga gtccctacac aatccatacc
     2041 ccgactacca ctggcttcga actgtaagaa ccaccaaaga gtccctcatt atcatatccc
     2101 caagtgtgac agatttggac ccatatgaca aatcccttca ctcaagggtc ttccctggcg
     2161 gaaagtgctc aggaataacg gtgtcctcta cctactgctc aactaaccat gattacacca
     2221 tttggatgcc cgagaatccg agaccaagga caccttgtga catttttacc aatagcagag
     2281 ggaagagagc atccaacggg aacaagactt gcggctttgt ggatgaaaga ggcctgtata
     2341 agtctctaaa aggagcatgc aggctcaagt tatgtggagt tcttggactt agacttatgg
     2401 atggaacatg ggtcgcgatg caaacatcag atgagaccaa atggtgccct ccagatcagt
     2461 tggtgaattt gcacgacttt cgctcagacg agattgagca tctcgttgtg gaggagttag
     2521 tcaagaaaag agaggaatgt ctggacgcat tagagtccat catgaccacc aagtcagtaa
     2581 gtttcagacg tctcagtcac ctgagaaaac ttgtcccagg gtttggaaaa gcatatacca
     2641 tattcaacaa aaccttgatg gaggctgatg ctcactacaa gtcagtccgg acctggaatg
     2701 agatcatccc ctcaaaaggg tgtttgaaag ttggaggaag gtgccatcct catgtgaacg
     2761 gggtgttttt caatggtata atattagggc ctgacgacca tgtcctaatc ccagagatgc
     2821 aatcatccct cctccagcaa catatggagt tgttggaatc ttcagttatc cccctgatgc
     2881 accccctggc agacccttct acagttttca aagaaggtga tgaggctgag gattttgttg
     2941 aagttcacct ccccgatgtg tacaaacaga tctcaggggt tgacctgggt ctcccgaact
     3001 ggggaaagta tgtattgatg actgcagggg ccatgattgg cctggtgttg atattttccc
     3061 taatgacatg gtgcagaaga gccaatcgac cagaatcgaa acaacgcagt tttggaggga
     3121 caggggggaa tgtgtcagtc acttcccaaa gcggaaaagt cataccttca tgggaatcat
     3181 ataagagtgg aggtgagatc agactgtaag atgtacaagt aaggcgcgcc acccctgcag
     3241 ggaattcgat atcaagctta tcgataatca acctctggat tacaaaattt gtgaaagatt
     3301 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     3361 tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg
     3421 gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac
     3481 tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc
     3541 cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc
     3601 ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa
     3661 atcatcgtcc tttccttggc tgctcgccta tgttgccacc tggattctgc gcgggacgtc
     3721 cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc
     3781 ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg
     3841 ggccgcctcc ccgcatcgat accgagcgct gctcgagaga tctgtgatag cggccatcaa
     3901 gctggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
     3961 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
     4021 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4081 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4141 cagcttatcg ataccgcatg cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg
     4201 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     4261 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc
     4321 aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     4381 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     4441 tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt
     4501 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     4561 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga
     4621 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     4681 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg
     4741 ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct
     4801 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg
     4861 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     4921 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     4981 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     5041 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     5101 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     5161 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     5221 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     5281 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     5341 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     5401 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     5461 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     5521 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     5581 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     5641 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     5701 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5761 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     5821 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     5881 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     5941 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     6001 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     6061 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     6121 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     6181 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     6241 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     6301 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     6361 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     6421 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     6481 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     6541 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     6601 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     6661 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     6721 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     6781 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     6841 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     6901 ctggcctttt gctcacatgt
//
