LOCUS       sequence-486012-        5619 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV transfer plasmid expressing eGFP-OLLAS under a CAG promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5619)
  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 5619)
  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-OLLAS
FEATURES             Location/Qualifiers
     source          1..5619
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(80..871,872..940))
                     /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     703..722
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(941..1045)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1071..1526)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1208..1229)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1420..1439
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   1598..1727
                     /label=AAV2 ITR
     promoter        2038..2315
                     /label=chicken beta-actin promoter
     primer_bind     2594..2616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2678..2697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2730..2739
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2736..2738,2739..2741,2742..3452)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2736..2738,2739..2741,2742..3452)
                     /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(2781..2802)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3042..3061)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3389..3410
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3471..3512
                     /codon_start=1
                     /product="Escherichia coli OmpF Linker and mouse Langerin 
                     fusion sequence tag (Park et al., 2008)"
                     /label=OLLAS tag
                     /translation="SGFANELGPRLMGK"
     CDS             3531..3572
                     /codon_start=1
                     /product="Escherichia coli OmpF Linker and mouse Langerin 
                     fusion sequence tag (Park et al., 2008)"
                     /label=OLLAS tag
                     /translation="SGFANELGPRLMGK"
     CDS             3591..3632
                     /codon_start=1
                     /product="Escherichia coli OmpF Linker and mouse Langerin 
                     fusion sequence tag (Park et al., 2008)"
                     /label=OLLAS tag
                     /translation="SGFANELGPRLMGK"
     misc_feature    3660..4248
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3713..3733)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4131..4142)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4291..4412
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4302..4321)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4356..4375
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4574..4703
                     /label=AAV2 ITR
     rep_origin      complement(4940..5528)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5020..5039)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
       61 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
      121 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
      181 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
      241 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
      301 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
      361 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
      421 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
      481 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
      541 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
      601 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
      661 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
      721 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
      781 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
      841 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
      901 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
      961 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     1021 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct aaattgtaag
     1081 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     1141 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     1201 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     1261 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     1321 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     1381 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     1441 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     1501 gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca actgttggga
     1561 agggcgatcg gtgcgggcct cttcgctatt acgccagctg cgcgctcgct cgctcactga
     1621 ggccgcccgg gcaaagcccg ggcgtcgggc gacctttggt cgcccggcct cagtgagcga
     1681 gcgagcgcgc agagagggag tggccaactc catcactagg ggttccttgt agttaatgat
     1741 taacccgcca tgctacttat ctacgtagcc atgctctagg aagagtacca ttgacgtcaa
     1801 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
     1861 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
     1921 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
     1981 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg
     2041 aggtgagccc cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt
     2101 tgtatttatt tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggggc
     2161 gcgcgccrgg cggggcgggg cggggcgrgg ggcggggcgg ggcgaggcgg agaggtgcgg
     2221 cggcagccaa tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc
     2281 ggcggcccta taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc
     2341 ccgtgccccg ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg accgcgttac
     2401 tcccacaggt gagcgggcgg gacggccctt ctcctccggg ctgtaattag cgcttggttt
     2461 aatgacggct tgtttctttt ctgtggctgc gtgaaagcct tgaggggctc cgggagggcc
     2521 ctttgtgcgg ggggagcggc tcggggctgt ccgcgggggg acggctgcct tcggggggga
     2581 cggggcaggg cggggttcgg cttctggcgt gtgaccggcg gctctagagc ctctgctaac
     2641 catgttcatg ccttcttctt tttcctacag ctcctgggca acgtgctggt tattgtgctg
     2701 tctcatcatt ttggcaaaga attggatccg ccaccatggt gagcaagggc gaggagctgt
     2761 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
     2821 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcattt
     2881 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
     2941 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
     3001 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
     3061 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     3121 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     3181 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     3241 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     3301 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga
     3361 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     3421 ggatcactct cggcatggac gagctgtaca aggggggatc aggcggaagc tctggatttg
     3481 ctaacgaact cggtccccga ctgatgggca aggggggaag tggaggttca agtggtttcg
     3541 ctaatgagtt gggaccacgc cttatgggga agggtggctc tggcgggagc tcaggatttg
     3601 caaatgagct cggtcccagg ttgatgggca aataagaatt cgatatcaag cttatcgata
     3661 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     3721 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     3781 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     3841 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     3901 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     3961 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     4021 tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg
     4081 cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     4141 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     4201 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgtc
     4261 gacccgggcg gccgcttcga gcagacatga taagatacat tgatgagttt ggacaaacca
     4321 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
     4381 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt
     4441 ttcaggttca gggggagatg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg
     4501 gtaaaatcga taaggatctt cctagagcat ggctacgtag ataagtagca tggcgggtta
     4561 atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc
     4621 tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc
     4681 tcagtgagcg agcgagcgcg cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     4741 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     4801 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     4861 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     4921 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     4981 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5041 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5101 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5161 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5221 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5281 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5341 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5401 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     5461 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5521 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5581 cacgttaagg gattttggtc atgagattat caaaaagga
//
