LOCUS       sequence-391644-        7083 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV constitutively expressing nuclear FLAG and Rabies Glycoprotein 
            (G) of CVS-N2c strain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7083)
  AUTHORS   Lee H, Ciabatti E, Gonzalez-Rueda A, Williams E, Nugent F, Mookerjee
            S, Morgese F, Tripodi M
  TITLE     Combining long-term circuit mapping and network transcriptomics with
            SiR-N2c.
  JOURNAL   Nat Methods. 2023 Apr;20(4):580-589. doi: 
            10.1038/s41592-023-01787-1. Epub 2023 Mar 2.
  PUBMED    36864202
REFERENCE   2  (bases 1 to 7083)
  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"; doi: 
            "10.1038/s41592-023-01787-1"; journalName: "Nat Methods"; date: 
            "2023-04"; volume: "20"; issue: "4"; pages: "580-589"
COMMENT     Sequence Label: pAAV-CMV-H2B-3xFLAG-2A-G(N2c)
FEATURES             Location/Qualifiers
     source          1..7083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   651..780
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        856..1159
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1160..1363
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1314..1334
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1360..1384
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          1512..1890
                     /label=chimeric intron
                     /note="fusion between human cytomegalovirus intron A and 
                     human beta-globin intron 2"
     CDS             1991..2368
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAVTKYTSAK"
     CDS             2453..2518
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             2528..2584
                     /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"
     primer_bind     complement(4190..4210)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4608..4619)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4760..4777)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4766..4973
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4996..5136
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4996..5125
                     /label=AAV2 ITR
     rep_origin      5211..5666
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5298..5317)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5508..5529
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5683..5702)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5802..5824
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5862..5880)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5948..6052
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6053..6121,6122..6913)
                     /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(6271..6290)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt cctgcaggca
      661 gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt ggtcgcccgg
      721 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      781 gcggccgcac gcgtctagtt attaatagta atcaattacg gggtcattag ttcatagccc
      841 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      901 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      961 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     1021 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     1081 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     1141 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
     1201 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
     1261 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
     1321 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag tgaaccgtca
     1381 gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc
     1441 cagcctccgc ggattcgaat cccggccggg aacggtgcat tggaacgcgg attccccgtg
     1501 ccaagagtga cgtaagtacc gcctatagag tctataggcc cacaaaaaat gctttcttct
     1561 tttaatatac ttttttgttt atcttatttc taatactttc cctaatctct ttctttcagg
     1621 gcaataatga tacaatgtat catgcctctt tgcaccattc taaagaataa cagtgataat
     1681 ttctgggtta aggcaatagc aatatttctg catataaata tttctgcata taaattgtaa
     1741 ctgatgtaag aggtttcata ttgctaatag cagctacaat ccagctacca ttctgctttt
     1801 attttatggt tgggataagg ctggattatt ctgagtccaa gctaggccct tttgctaatc
     1861 atgttcatac ctcttatctt cctcccacag ctcctgggca acgtgctggt ctgtgtgctg
     1921 gcccatcact ttggcaaaga attgggattc gaacatcgat tgaattcggt acccttcagg
     1981 tcgacccacc atgccagagc cagcgaagtc tgctcccgcc ccgaaaaagg gctccaagaa
     2041 ggcggtgact aaggcgcaga agaaaggcgg caagaagcgc aagcgcagcc gcaaggagag
     2101 ctattccatc tatgtgtaca aggttctgaa gcaggtccac cctgacaccg gcatttcgtc
     2161 caaggccatg ggcatcatga actcgtttgt gaacgacatt ttcgagcgca tcgcaggtga
     2221 ggcttcccgc ctggcgcatt acaacaagcg ctcgaccatc acctccaggg agatccagac
     2281 ggccgtgcgc ctgctgctgc ctggggagtt ggccaagcac gccgtgtccg agggtactaa
     2341 ggccgtcacc aagtacacca gcgctaaggg atctgggggt ggaggctccg gcgggggtgg
     2401 atctggaggt gggggcagcg gcggaggtgg gctagcgcta ccgcgggcca ccgactacaa
     2461 agaccatgac ggtgattata aagatcatga catcgactac aaggatgacg atgacaaggg
     2521 atcgggagcc acgaacttct ctctgttaaa gcaagcagga gacgtggaag aaaaccccgg
     2581 tcctatggtg cctcaggtgc tgctgtttgt gctgctgctg ggcttcagcc tgtgcttcgg
     2641 caagttcccc atctacacca tccccgacga gctgggccct tggagcccta tcgatatcca
     2701 ccacctgagc tgccccaaca acctggtggt ggaagatgag ggctgcacca acctgagcga
     2761 gttcagctac atggaactga aagtgggcta catcagcgcc atcaaagtga acggcttcac
     2821 ctgtaccggc gtcgtgaccg aggccgagac atacaccaac ttcgtgggct acgtgaccac
     2881 caccttcaag agaaagcact tcagacccac ccccgacgcc tgcagagccg cctacaattg
     2941 gaagatggcc ggcgacccca gatacgagga aagcctgcac aacccctacc ccgactacca
     3001 ctggctgaga accgtgcgga ccaccaaaga gagcctgatc atcatcagcc ccagcgtgac
     3061 cgacctggac ccctacgaca agagcctgca ctccagagtg ttccctggcg gcaagtgcag
     3121 cggcatcacc gtgtctagca cctactgcag caccaaccac gactacacaa tctggatgcc
     3181 cgagaacccc agacccagaa ccccctgcga catcttcacc aacagcagag gcaagagagc
     3241 cagcaacggc aacaagacct gcggcttcgt ggacgagaga ggcctgtaca agtccctgaa
     3301 gggcgcctgc cggctgaagc tgtgtggcgt gctgggactg agactgatgg acggcacatg
     3361 ggtggccatg cagaccagcg acgagacaaa gtggtgcccc cctgatcagc tcgtgaacct
     3421 gcacgacttc agatccgacg agatcgagca tctggtggtg gaagaactcg tgaagaaaag
     3481 ggaagagtgc ctggacgccc tggaatccat catgaccacc aagagcgtgt ccttcagaag
     3541 gctgagccac ctgagaaagc tggtgcccgg ctttggcaag gcttacacca tcttcaacaa
     3601 gacactgatg gaagccgacg cccactacaa gtctgtgcgg acctggaacg agatcatccc
     3661 cagcaagggc tgcctgaaag tgggaggcag atgccaccct cacgtgaacg gcgtgttctt
     3721 caacggcatc atcctgggcc ccgacgacca cgtgctgatc cctgagatgc agagcagcct
     3781 gctgcagcag catatggaac tgctggaaag cagcgtgatc cccctgatgc accctctggc
     3841 cgaccctagc accgtgttca aagagggcga cgaggctgag gacttcgtgg aagtgcatct
     3901 gcccgacgtg tacaagcaga tcagcggcgt ggacctgggc ctgcccaact ggggcaaata
     3961 tgtgctgatg accgctggcg ccatgatcgg cctggtgctg atcttctccc tgatgacctg
     4021 gtgcagaagg gccaacagac ccgagagcaa gcagagaagc ttcggcggca caggcggcaa
     4081 cgtgtccgtg acaagccaga gcggcaaagt gatccctagc tgggagagct acaagagcgg
     4141 cggagagatc agactgtgat gatgaaagct tgcctggtat tcttaactat gttgctcctt
     4201 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     4261 ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc
     4321 ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt
     4381 ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg
     4441 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
     4501 gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct
     4561 atgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc
     4621 cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc
     4681 ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgagcgc
     4741 tgctcgagag atcgatctgc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     4801 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     4861 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     4921 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggacacgtgc
     4981 ggaccgagcg gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
     5041 gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg
     5101 cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc
     5161 ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg
     5221 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     5281 cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     5341 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     5401 gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg
     5461 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     5521 ccaaactgga acaacactca actctatctc gggctattct tttgatttat aagggatttt
     5581 gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     5641 taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc
     5701 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     5761 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     5821 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     5881 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     5941 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     6001 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     6061 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     6121 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     6181 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     6241 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     6301 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     6361 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     6421 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     6481 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     6541 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     6601 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     6661 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     6721 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     6781 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     6841 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     6901 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     6961 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     7021 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     7081 ttc
//
