LOCUS       Exported                6270 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Plasmid name in publication: pAAV-ER-dAPEX2. Peroxidase reporter for
            multiplexed electron microscopy labeling.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-IGK-dAPEX2-KDEL
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6270)
  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 6270)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6270
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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    1440..1473
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites (Lee and 
                     Saito, 1988)."
     regulatory      1484..1493
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     sig_peptide     1490..1552
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
     CDS             1553..2299
                     /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"
     protein_bind    complement(2339..2372)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    2397..2985
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2450..2470)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2868..2879)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3017..3493
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3216..3235)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3533..3673
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3533..3662
                     /label=AAV2 ITR
     rep_origin      3748..4203
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3835..3854)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4045..4066
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4220..4239)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4339..4361
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4399..4417)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4485..4589
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4590..5450
                     /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(4808..4827)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5621..6209
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6110..6129
                     /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 tatgctagaa ttcgccacca tggagacaga
     1501 cacactcctg ctatgggtac tgctgctctg ggttccaggt tccactggtg acggaaagtc
     1561 ttacccaact gtgagtgctg attaccagaa ggccgttgag aaggcgaaga agaagctcag
     1621 aggcttcatc gctgagaaga gatgcgctcc tctaatgctc cgtttggcat tccactctgc
     1681 tggaaccttt gacaagggca cgaagaccgg tggacccttc ggaaccatca agcaccctgc
     1741 cgaactggct cacagcgcta acaacggtct tgacatcgct gttaggcttt tggagccact
     1801 caaggcggag ttccctattt tgagctacgc cgatttctac cagttggctg gcgttgttgc
     1861 cgttgaggtc acgggtggac ctgaagttcc attccaccct ggaagagagg acaagcctga
     1921 gccaccacca gagggtcgct tgcccgatcc cactaagggt tctgaccatt tgagagatgt
     1981 gtttggcaaa gctatggggc ttactgacca agatatcgtt gctctatctg ggggtcacac
     2041 tattggagct gcacacaagg agcgttctgg atttgagggt ccctggacct ctaatcctct
     2101 tattttcgac aactcatact tcacggagtt gttgagtggt gagaaggaag gtctccttca
     2161 gctaccttct gacaaggctc ttttgtctga ccctgtattc cgccctctcg ttgataaata
     2221 tgcagcggac gaagatgcct tctttgctga ttacgctgag gctcaccaaa agctttccga
     2281 gcttgggttt gctgatgcca aggacgagct gtaagctagc gtacaagtaa ggcgcgccat
     2341 aacttcgtat aatgtatgct atacgaagtt atgaattcga tatcaagctt atcgataatc
     2401 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     2461 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     2521 ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc
     2581 ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt
     2641 ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg
     2701 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
     2761 gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct
     2821 atgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc
     2881 cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc
     2941 ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgagcgc
     3001 tgctcgagag atctacgggt ggcatccctg tgacccctcc ccagtgcctc tcctggccct
     3061 ggaagttgcc actccagtgc ccaccagcct tgtcctaata aaattaagtt gcatcatttt
     3121 gtctgactag gtgtccttct ataatattat ggggtggagg ggggtggtat ggagcaaggg
     3181 gcaagttggg aagacaacct gtagggcctg cggggtctat tgggaaccaa gctggagtgc
     3241 agtggcacaa tcttggctca ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc
     3301 tcagcctccc gagttgttgg gattccaggc atgcatgacc aggctcagct aatttttgtt
     3361 tttttggtag agacggggtt tcaccatatt ggccaggctg gtctccaact cctaatctca
     3421 ggtgatctac ccaccttggc ctcccaaatt gctgggatta caggcgtgaa ccactgctcc
     3481 cttccctgtc cttctgattt tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc
     3541 ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga
     3601 ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc
     3661 agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     3721 caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg
     3781 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt
     3841 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     3901 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     3961 atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     4021 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact
     4081 ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa
     4141 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa
     4201 ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac
     4261 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     4321 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     4381 aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     4441 taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     4501 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     4561 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     4621 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     4681 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     4741 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     4801 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     4861 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     4921 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     4981 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     5041 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     5101 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     5161 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     5221 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     5281 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     5341 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     5401 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     5461 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     5521 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     5581 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     5641 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     5701 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     5761 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     5821 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     5881 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     5941 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     6001 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     6061 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     6121 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     6181 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     6241 tggccttttg ctggcctttt gctcacatgt
//
