LOCUS       sequence-225124-        6438 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid name in publication: pAAV-DIO-ER-dAPEX2. Peroxidase reporter
            for multiplexed electron microscopy labeling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6438)
  AUTHORS   Zhang Q, Lee WA, Paul DL, Ginty DD
  TITLE     Multiplexed peroxidase-based electron microscopy labeling enables 
            simultaneous visualization of multiple cell types.
  JOURNAL   Nat Neurosci. 2019 Mar 18. pii: 10.1038/s41593-019-0358-7. doi: 
            10.1038/s41593-019-0358-7.
  PUBMED    30886406
REFERENCE   2  (bases 1 to 6438)
  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: "Nat 
            Neurosci. 2019 Mar 18. pii: 10.1038/s41593-019-0358-7. doi: 
            10.1038/s41593-019-0358-7."
COMMENT     Sequence Label: pAAV-DIO-IGK-dAPEX2-KDEL
FEATURES             Location/Qualifiers
     source          1..6438
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1440..1473)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1524..1557)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1597..2343)
                     /codon_start=1
                     /product="soybean ascorbate peroxidase, engineered to serve
                     as a monomeric tag for electron microscopy and live-cell 
                     proteomics (Lam et al., 2015)"
                     /label=APEX2
                     /note="more active than the original APEX due to the A134P 
                     mutation"
                     /translation="GKSYPTVSADYQKAVEKAKKKLRGFIAEKRCAPLMLRLAFHSAGT
                     FDKGTKTGGPFGTIKHPAELAHSANNGLDIAVRLLEPLKAEFPILSYADFYQLAGVVAV
                     EVTGGPEVPFHPGREDKPEPPPEGRLPDPTKGSDHLRDVFGKAMGLTDQDIVALSGGHT
                     IGAAHKERSGFEGPWTSNPLIFDNSYFTELLSGEKEGLLQLPSDKALLSDPVFRPLVDK
                     YAADEDAFFADYAEAHQKLSELGFADA"
     sig_peptide     complement(2347..2406)
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     regulatory      2403..2412
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2423..2456
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    2507..2540
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2565..3153
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2618..2638)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3036..3047)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3185..3661
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3384..3403)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3701..3841
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3701..3830
                     /label=AAV2 ITR
     rep_origin      3916..4371
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4003..4022)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4213..4234
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4388..4407)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4507..4529
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4567..4585)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4653..4757
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4758..4826,4827..5618)
                     /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(4976..4995)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5789..6377
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6278..6297
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggaa
     1441 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
     1561 gcgccttact tgtacgctag cttacagctc gtccttggca tcagcaaacc caagctcgga
     1621 aagcttttgg tgagcctcag cgtaatcagc aaagaaggca tcttcgtccg ctgcatattt
     1681 atcaacgaga gggcggaata cagggtcaga caaaagagcc ttgtcagaag gtagctgaag
     1741 gagaccttcc ttctcaccac tcaacaactc cgtgaagtat gagttgtcga aaataagagg
     1801 attagaggtc cagggaccct caaatccaga acgctccttg tgtgcagctc caatagtgtg
     1861 acccccagat agagcaacga tatcttggtc agtaagcccc atagctttgc caaacacatc
     1921 tctcaaatgg tcagaaccct tagtgggatc gggcaagcga ccctctggtg gtggctcagg
     1981 cttgtcctct cttccagggt ggaatggaac ttcaggtcca cccgtgacct caacggcaac
     2041 aacgccagcc aactggtaga aatcggcgta gctcaaaata gggaactccg ccttgagtgg
     2101 ctccaaaagc ctaacagcga tgtcaagacc gttgttagcg ctgtgagcca gttcggcagg
     2161 gtgcttgatg gttccgaagg gtccaccggt cttcgtgccc ttgtcaaagg ttccagcaga
     2221 gtggaatgcc aaacggagca ttagaggagc gcatctcttc tcagcgatga agcctctgag
     2281 cttcttcttc gccttctcaa cggccttctg gtaatcagca ctcacagttg ggtaagactt
     2341 tccgtcacca gtggaacctg gaacccagag cagcagtacc catagcagga gtgtgtctgt
     2401 ctccatggtg gcgaattcta gcataacttc gtataaagta tcctatacga agttatttgc
     2461 cttaacccag aaattatcac tgttattctt tagaatggtg caaagaataa cttcgtataa
     2521 tgtatgctat acgaagttat gaattcgata tcaagcttat cgataatcaa cctctggatt
     2581 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     2641 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     2701 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     2761 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     2821 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     2881 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     2941 ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctat gttgccacct
     3001 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     3061 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     3121 cgagtcggat ctccctttgg gccgcctccc cgcatcgata ccgagcgctg ctcgagagat
     3181 ctacgggtgg catccctgtg acccctcccc agtgcctctc ctggccctgg aagttgccac
     3241 tccagtgccc accagccttg tcctaataaa attaagttgc atcattttgt ctgactaggt
     3301 gtccttctat aatattatgg ggtggagggg ggtggtatgg agcaaggggc aagttgggaa
     3361 gacaacctgt agggcctgcg gggtctattg ggaaccaagc tggagtgcag tggcacaatc
     3421 ttggctcact gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc agcctcccga
     3481 gttgttggga ttccaggcat gcatgaccag gctcagctaa tttttgtttt tttggtagag
     3541 acggggtttc accatattgg ccaggctggt ctccaactcc taatctcagg tgatctaccc
     3601 accttggcct cccaaattgc tgggattaca ggcgtgaacc actgctccct tccctgtcct
     3661 tctgattttg taggtaacca cgtgcggacc gagcggccgc aggaacccct agtgatggag
     3721 ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc
     3781 cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag
     3841 gggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt
     3901 caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     3961 cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc
     4021 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     4081 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg
     4141 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     4201 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct
     4261 attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga
     4321 tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca
     4381 ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac
     4441 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     4501 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac
     4561 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
     4621 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     4681 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     4741 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     4801 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     4861 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     4921 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     4981 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     5041 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     5101 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     5161 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     5221 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     5281 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     5341 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     5401 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     5461 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     5521 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     5581 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     5641 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     5701 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     5761 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     5821 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     5881 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     5941 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     6001 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     6061 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     6121 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     6181 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     6241 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     6301 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     6361 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     6421 ggccttttgc tcacatgt
//
