LOCUS       sequence-403655-        4977 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For barcoded retrograde labeling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4977)
  AUTHORS   Xu P, Peng J, Yuan T, Chen Z, He H, Wu Z, Li T, Li X, Wang L, Gao L,
            Yan J, Wei W, Li CT, Luo ZG, Chen Y
  TITLE     High-throughput mapping of single-neuron projection and molecular 
            features by retrograde barcoded labeling.
  JOURNAL   Elife. 2024 Feb 23;13:e85419. doi: 10.7554/eLife.85419.
  PUBMED    38390967
REFERENCE   2  (bases 1 to 4977)
  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: "2024-02-23"; volume: "13:e85419. doi"; pages: " 
            10.7554/eLife.85419"
COMMENT     Sequence Label: pAAV-CAG-EGFP barcode-9-SV40 polyA
FEATURES             Location/Qualifiers
     source          1..4977
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        414..690
                     /label=chicken beta-actin promoter
     primer_bind     969..991
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1053..1072
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1118..1127
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1124..1126,1127..1129,1130..1843)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1124..1126,1127..1129,1130..1840)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1169..1190)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1430..1449)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1777..1798
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1913..1930)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1919..2143
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2308..2437
                     /label=AAV2 ITR
     rep_origin      complement(2674..3262)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2754..2773)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3433..4224,4225..4293))
                     /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     4056..4075
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4294..4398)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4424..4879)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4561..4582)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4773..4792
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   join(4951..4977,1..103)
                     /label=AAV2 ITR
ORIGIN
        1 gcccgggcaa agcccgggcg tcgggcgacc tttggtcgcc cggcctcagt gagcgagcga
       61 gcgcgcagag agggagtggc caactccatc actaggggtt ccttgtagtt aatgattaac
      121 ccgccatgct acttatctac gtagccatgc tctaggaaga gtaccattga cgtcaataat
      181 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      241 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
      301 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
      361 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtcgaggt
      421 gagccccacg ttctgcttca ctctccccat ctcccccccc tccccacccc caattttgta
      481 tttatttatt ttttaattat tttgtgcagc gatgggggcg gggggggggg ggrscgcgcg
      541 ccaggcgggg cggggcgggg cgaggggcgg ggcggggcga ggcggagagg tgcggcggca
      601 gccaatcaga gcggcgcgct ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg
      661 ccctataaaa agcgaagcgc gcggcgggcg ggagtcgctg cgcgctgcct tcgccccgtg
      721 ccccgctccg ccgccgcctc gcgccgcccg ccccggctct gactgaccgc gttactccca
      781 caggtgagcg ggcgggacgg cccttctcct ccgggctgta attagcgctt ggtttaatga
      841 cggcttgttt cttttctgtg gctgcgtgaa agccttgagg ggctccggga gggccctttg
      901 tgcgggggga gcggctcggg gctgtccgcg gggggacggc tgccttcggg ggggacgggg
      961 cagggcgggg ttcggcttct ggcgtgtgac cggcggctct agagcctctg ctaaccatgt
     1021 tcatgccttc ttctttttcc tacagctcct gggcaacgtg ctggttattg tgctgtctca
     1081 tcattttggc aaagaattgg atccccgggt accggtcgcc accatggtga gcaagggcga
     1141 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     1201 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     1261 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
     1321 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
     1381 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     1441 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     1501 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     1561 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
     1621 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
     1681 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
     1741 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     1801 cgccgccggg atcactctcg gcatggacga gctgtacaag taaagcggcc gcgactctag
     1861 aattcctgtg ccatcggcat ggatccaggg cccgtttaaa cccgctgatc agcctcgact
     1921 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     1981 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     2041 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     2101 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tgggttaaca acaacaattg
     2161 cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa gcaagtaaaa
     2221 cctctacaaa tgtggtaaaa tcgataagga tcttcctaga gcatggctac gtagataagt
     2281 agcatggcgg gttaatcatt aactacaagg aacccctagt gatggagttg gccactccct
     2341 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct
     2401 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cattaatgaa tcggccaacg
     2461 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     2521 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     2581 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     2641 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     2701 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2761 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2821 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     2881 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2941 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3001 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3061 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     3121 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3181 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3241 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3301 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3361 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3421 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3481 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     3541 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     3601 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     3661 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     3721 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     3781 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3841 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     3901 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     3961 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4021 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4081 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4141 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4201 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4261 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4321 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4381 acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat
     4441 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     4501 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt
     4561 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa
     4621 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg
     4681 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     4741 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     4801 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     4861 gcgccgctac agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     4921 tcggtgcggg cctcttcgct attacgccag ctgcgcgctc gctcgctcac tgaggcc
//
