LOCUS       sequence-51844-e        6390 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6390)
  AUTHORS   Martell JD, Deerinck TJ, Sancak Y, Poulos TL, Mootha VK, Sosinsky 
            GE, Ellisman MH, Ting AY
  TITLE     Engineered ascorbate peroxidase as a genetically encoded reporter 
            for electron microscopy.
  JOURNAL   Nat Biotechnol. 2012 Oct 21. doi: 10.1038/nbt.2375.
  PUBMED    23086203
REFERENCE   2  (bases 1 to 6390)
  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 
            Biotechnol. 2012 Oct 21. doi: 10.1038/nbt.2375."
COMMENT     Sequence Label: pcDNA3 soybean APEX (IMS)
FEATURES             Location/Qualifiers
     source          1..6390
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1120..1161
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             1162..1908
                     /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="GKSYPTVSADYQDAVEKAKKKLRGFIAEKRCAPLMLRLAFHSAGT
                     FDKGTKTGGPFGTIKHPAELAHSANNGLDIAVRLLEPLKAEFPILSYADFYQLAGVVAV
                     EVTGGPKVPFHPGREDKPEPPPEGRLPDATKGSDHLRDVFGKAMGLTDQDIVALSGGHT
                     IGAAHKERSGFEGPWTSNPLIFDNSYFTELLSGEKEGLLQLPSDKALLSDPVFRPLVDK
                     YAADEDAFFADYAEAHQKLSELGFADA"
     promoter        complement(1942..1960)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1943..1960)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1980..1997)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1986..2210
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2256..2684
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2343..2362)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2553..2574
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2693..2713)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2698..3028
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2879..3014
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2941..2960
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3095..3889
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3149..3168)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3759..3778
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4063..4184
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4100..4119)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4154..4173
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4233..4249)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4233..4249)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4246..4268)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4257..4273
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4281..4287,4288..4305,4306..4311))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4326..4347
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4464..4481)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4635..5223)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4715..4734)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5394..6185,6186..6254))
                     /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     6017..6036
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6255..6359)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgt accggctcct gtcaagcgtg acagctcggg ctgcggccac cgcaggccca
      961 gcctgggacg gagggcggcg cggggcgcac aggcgaccgg gcctgcctgt gctgggcctt
     1021 ggttgggccg gcggcctggg gctcgggctg gggctggctc tcggcgcgaa gctggtggtc
     1081 gggctgcggg gcgccgtccc cattcagtcc agggatccag gcaagcccat ccccaacccc
     1141 ctgctgggcc tggacagcac cggaaagtct tacccaactg tgagtgctga ttaccaggac
     1201 gccgttgaga aggcgaagaa gaagctcaga ggcttcatcg ctgagaagag atgcgctcct
     1261 ctaatgctcc gtttggcatt ccactctgct ggaacctttg acaagggcac gaagaccggt
     1321 ggacccttcg gaaccatcaa gcaccctgcc gaactggctc acagcgctaa caacggtctt
     1381 gacatcgctg ttaggctttt ggagccactc aaggcggagt tccctatttt gagctacgcc
     1441 gatttctacc agttggctgg cgttgttgcc gttgaggtca cgggtggacc taaggttcca
     1501 ttccaccctg gaagagagga caagcctgag ccaccaccag agggtcgctt gcccgatgcc
     1561 actaagggtt ctgaccattt gagagatgtg tttggcaaag ctatggggct tactgaccaa
     1621 gatatcgttg ctctatctgg gggtcacact attggagctg cacacaagga gcgttctgga
     1681 tttgagggtc cctggacctc taatcctctt attttcgaca actcatactt cacggagttg
     1741 ttgagtggtg agaaggaagg tctccttcag ctaccttctg acaaggctct tttgtctgac
     1801 cctgtattcc gccctctcgt tgataaatat gcagcggacg aagatgcctt ctttgctgat
     1861 tacgctgagg ctcaccaaaa gctttccgag cttgggtttg ctgatgccta atgactcgag
     1921 catgcatcta gagggcccta ttctatagtg tcacctaaat gctagagctc gctgatcagc
     1981 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     2041 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     2101 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     2161 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     2221 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     2281 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     2341 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     2401 tctaaatcgg ggcatccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     2461 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     2521 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     2581 actcaaccct atctcggtct attcttttga tttataaggg attttgggga tttcggccta
     2641 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     2701 tgtcagttag ggtgtggaaa gtccccaggc tccccaggca ggcagaagta tgcaaagcat
     2761 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     2821 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     2881 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     2941 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg
     3001 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg
     3061 atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     3121 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     3181 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     3241 caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg
     3301 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     3361 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     3421 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     3481 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     3541 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     3601 actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg
     3661 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     3721 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     3781 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     3841 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     3901 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     3961 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     4021 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     4081 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     4141 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     4201 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     4261 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     4321 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4381 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4441 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4501 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4561 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4621 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4681 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4741 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4801 ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct gtaggtatct cagttcggtg
     4861 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4921 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4981 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     5041 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     5101 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     5161 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     5221 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     5281 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     5341 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     5401 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     5461 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     5521 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     5581 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     5641 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     5701 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     5761 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     5821 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     5881 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     5941 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     6001 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     6061 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     6121 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     6181 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     6241 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     6301 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     6361 acatttcccc gaaaagtgcc acctgacgtc
//
