LOCUS       sequence-486028-        5572 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV transfer plasmid expressing eGFP-HA under a CAG promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5572)
  AUTHORS   Park SY, Sheridan A, An B, Jarvis E, Lyudchik J, Patton W, Axup JY, 
            Chan SW, Damstra HGJ, Leible D, Leung KS, Magno CA, Meeran A, 
            Michalska JM, Rieger F, Wang C, Wu M, Church GM, Funke J, Huffman T,
            Leeper KGC, Truckenbrodt S, Winnubst J, Kornfeld JMR, Boyden ES, 
            Rodriques SG, Payne AC
  TITLE     Combinatorial protein barcodes enable self-correcting neuron tracing
            with nanoscale molecular context
  JOURNAL   bioRxiv 2025.09.26.678648
REFERENCE   2  (bases 1 to 5572)
  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: "bioRxiv 
            2025.09.26.678648"
COMMENT     Sequence Label: pAAV-CAG-eGFP-HA
FEATURES             Location/Qualifiers
     source          1..5572
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     14..33
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(252..356)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(382..837)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(519..540)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     731..750
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   909..1038
                     /label=AAV2 ITR
     promoter        1349..1624
                     /label=chicken beta-actin promoter
     primer_bind     1903..1925
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1987..2006
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2039..2048
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2045..2047,2048..2050,2051..2761)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2045..2047,2048..2050,2051..2761)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2090..2111)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2351..2370)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2698..2719
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2780..2806
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2825..2851
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2870..2896
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     misc_feature    2924..3512
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2977..2997)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3395..3406)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3555..3676
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3566..3585)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3620..3639
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3838..3967
                     /label=AAV2 ITR
     rep_origin      complement(4204..4792)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4284..4303)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4963..5572,1..182,183..251))
                     /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"
ORIGIN
        1 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
       61 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      121 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      181 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
      241 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
      301 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
      361 tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa
      421 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
      481 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
      541 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
      601 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
      661 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
      721 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
      781 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc
      841 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      901 tacgccagct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg
      961 cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact
     1021 ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc
     1081 catgctctag gaagagtacc attgacgtca ataatgacgt atgttcccat agtaacgcca
     1141 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
     1201 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
     1261 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
     1321 tacgtattag tcatcgctat taccatggtc gaggtgagcc ccacgttctg cttcactctc
     1381 cccatctccc ccccctcccc acccccaatt ttgtatttat ttatttttta attattttgt
     1441 gcagcgatgg gggcgggggg gggggggggg cgcgccaggc ggggcggggc ggggcgaggg
     1501 gcggggcggg gcgaggcgga gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa
     1561 gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg
     1621 ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg
     1681 cccgccccgg ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc
     1741 tcctccgggc tgtaattagc gcttggttta atgacggctt gtttcttttc tgtggctgcg
     1801 tgaaagcctt gaggggctcc gggagggccc tttgtgcggg gggagcggct cggggctgtc
     1861 cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg
     1921 tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     1981 tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttggatccgc
     2041 caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     2101 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     2161 ctacggcaag ctgaccctga agttcatttg caccaccggc aagctgcccg tgccctggcc
     2221 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     2281 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     2341 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     2401 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     2461 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     2521 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     2581 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     2641 ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     2701 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     2761 gggcgggagt ggtggaagtt atccatacga cgttcccgac tatgctggag gttccggcgg
     2821 atcttatccc tatgatgttc cagattacgc cggaggatct ggtggcagtt acccctatga
     2881 tgtgcccgac tacgcctaag aattcgatat caagcttatc gataatcaac ctctggatta
     2941 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     3001 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     3061 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     3121 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     3181 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     3241 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     3301 cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg
     3361 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     3421 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     3481 gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgtcgacccg ggcggccgct
     3541 tcgagcagac atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg
     3601 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     3661 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     3721 gatgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga
     3781 tcttcctaga gcatggctac gtagataagt agcatggcgg gttaatcatt aactacaagg
     3841 aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg
     3901 ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag
     3961 cgcgcagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     4021 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     4081 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     4141 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     4201 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4261 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4321 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4381 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4441 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4501 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4561 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4621 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4681 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4741 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4801 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     4861 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     4921 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     4981 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     5041 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     5101 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     5161 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     5221 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5281 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5341 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5401 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5461 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5521 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cg
//
