LOCUS       sequence-486995-        5600 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV transfer plasmid expressing eGFP-ProteinC under a CAG promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5600)
  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 5600)
  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-ProteinC
FEATURES             Location/Qualifiers
     source          1..5600
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      657..1245
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1146..1165
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1482..1611
                     /label=AAV2 ITR
     polyA_signal    1773..1894
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1810..1829)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1864..1883
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    1937..2525
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             2043..2054
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     2452..2472
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2553..2588)
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             complement(2607..2642)
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             complement(2661..2696)
                     /codon_start=1
                     /product="epitope tag derived from human Protein C (Stearns
                     et al., 1988)"
                     /label=Protein C tag
                     /translation="EDQVDPRLIDGK"
     CDS             complement(join(2715..3425,3426..3428,3429..3431))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(2715..3425,3426..3428,3429..3431))
                     /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(2757..2778)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3106..3125
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3365..3386
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3428..3437
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3470..3489)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(3551..3573)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     repeat_region   4439..4568
                     /label=AAV2 ITR
     rep_origin      4640..5095
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4727..4746)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4937..4958
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5121..5225
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5226..5294,5295..5600,1..486)
                     /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(5444..5463)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
       61 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
      121 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
      181 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
      241 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
      301 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
      361 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
      421 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
      481 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
      541 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
      601 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
      661 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
      721 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
      781 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
      841 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
      901 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
      961 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     1021 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     1081 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     1141 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     1201 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     1261 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     1321 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     1381 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     1441 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctgcgcgct cgctcgctca
     1501 ctgaggccgc ccgggcaaag cccgggcgtc gggcgacctt tggtcgcccg gcctcagtga
     1561 gcgagcgagc gcgcagagag ggagtggcca actccatcac taggggttcc ttgtagttaa
     1621 tgattaaccc gccatgctac ttatctacgt agccatgctc taggaagatc cttatcgatt
     1681 ttaccacatt tgtagaggtt ttacttgctt taaaaaacct cccacatctc cccctgaacc
     1741 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt
     1801 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     1861 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgctcgaag cggccgcccg
     1921 ggtcgacggt atcgatgcgg ggaggcggcc caaagggaga tccgactcgt ctgagggcga
     1981 aggcgaagac gcggaagagg ccgcagagcc ggcagcaggc cgcgggaagg aaggtccgct
     2041 ggattgaggg ccgaagggac gtagcagaag gacgtcccgc gcagaatcca ggtggcaaca
     2101 caggcgagca gccaaggaaa ggacgatgat ttccccgaca acaccacgga attgtcagtg
     2161 cccaacagcc gagcccctgt ccagcagcgg gcaaggcagg cggcgatgag ttccgccgtg
     2221 gcaataggga gggggaaagc gaaagtcccg gaaaggagct gacaggtggt ggcaatgccc
     2281 caaccagtgg gggttgcgtc agcaaacaca gtgcacacca cgccacgttg cctgacaacg
     2341 ggccacaact cctcataaag agacagcaac caggatttat acaaggagga gaaaatgaaa
     2401 gccatacggg aagcaatagc atgatacaaa ggcattaaag cagcgtatcc acatagcgta
     2461 aaaggagcaa catagttaag aataccagtc aatctttcac aaattttgta atccagaggt
     2521 tgattatcga taagcttgat atcgaattct tattttccgt caataagtcg tggatcaact
     2581 tggtcctcgc tccctccgct tccaccttta ccgtctatga gtcgtgggtc cacctggtct
     2641 tctgacccac ctgatccgcc cttgccatca atcaatcttg ggtcaacttg atcctcggac
     2701 cccccgctcc cacccttgta cagctcgtcc atgccgagag tgatcccggc ggcggtcacg
     2761 aactccagca ggaccatgtg atcgcgcttc tcgttggggt ctttgctcag ggcggactgg
     2821 gtgctcaggt agtggttgtc gggcagcagc acggggccgt cgccgatggg ggtgttctgc
     2881 tggtagtggt cggcgagctg cacgctgccg tcctcgatgt tgtggcggat cttgaagttc
     2941 accttgatgc cgttcttctg cttgtcggcc atgatataga cgttgtggct gttgtagttg
     3001 tactccagct tgtgccccag gatgttgccg tcctccttga agtcgatgcc cttcagctcg
     3061 atgcggttca ccagggtgtc gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg
     3121 tcgtccttga agaagatggt gcgctcctgg acgtagcctt cgggcatggc ggacttgaag
     3181 aagtcgtgct gcttcatgtg gtcggggtag cggctgaagc actgcacgcc gtaggtcagg
     3241 gtggtcacga gggtgggcca gggcacgggc agcttgccgg tggtgcaaat gaacttcagg
     3301 gtcagcttgc cgtaggtggc atcgccctcg ccctcgccgg acacgctgaa cttgtggccg
     3361 tttacgtcgc cgtccagctc gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc
     3421 ttgctcacca tggtggcgga tccaattctt tgccaaaatg atgagacagc acaataacca
     3481 gcacgttgcc caggagctgt aggaaaaaga agaaggcatg aacatggtta gcagaggctc
     3541 tagagccgcc ggtcacacgc cagaagccga accccgccct gccccgtccc ccccgaaggc
     3601 agccgtcccc ccgcggacag ccccgagccg ctccccccgc acaaagggcc ctcccggagc
     3661 ccctcaaggc tttcacgcag ccacagaaaa gaaacaagcc gtcattaaac caagcgctaa
     3721 ttacagcccg gaggagaagg gccgtcccgc ccgctcacct gtgggagtaa cgcggtcagt
     3781 cagagccggg gcgggcggcg cgaggcggcg gcggagcggg gcacggggcg aaggcagcgc
     3841 gcagcgactc ccgcccgccg cgcgcttcgc tttttatagg gccgccgccg ccgccgcctc
     3901 gccataaaag gaaactttcg gagcgcgccg ctctgattgg ctgccgccgc mccycyccsc
     3961 cycscccccs ccccsccccc cscccccccc cccccccccy ggcgcscscc cccccccccc
     4021 cngcccccat cgctgcacaa aataattaaa aaataaataa atacaaaatt gggggtgggg
     4081 agggggggga gatggggaga gtgaagcaga acgtggggct cacctcgacc atggtaatag
     4141 cgatgactaa tacgtagatg tactgccaag taggaaagtc ccataaggtc atgtactggg
     4201 cataatgcca ggcgggccat ttaccgtcat tgacgtcaat agggggcgta cttggcatat
     4261 gatacacttg atgtactgcc aagtgggcag tttaccgtaa atactccacc cattgacgtc
     4321 aatggaaagt ccctattggc gttactatgg gaacatacgt cattattgac gtcaatggta
     4381 ctcttcctag agcatggcta cgtagataag tagcatggcg ggttaatcat taactacaag
     4441 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc
     4501 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga
     4561 gcgcgcagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     4621 agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4681 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4741 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4801 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     4861 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     4921 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     4981 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     5041 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
     5101 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5161 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5221 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5281 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5341 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5401 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5461 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5521 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5581 aattatgcag tgctgccata
//
