LOCUS       sequence-326701-        6425 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Apex2 fused to Venus-CAAX for targeting to plasma 
            membrane. Useful for correlated light-eletron microscopy experiments
            in neurons..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6425)
  AUTHORS   Hirabayashi Y, Tapia JC, Polleux F
  TITLE     Correlated Light-Serial Scanning Electron Microscopy (CoLSSEM) for 
            ultrastructural visualization of single neurons in vivo.
  JOURNAL   Sci Rep. 2018 Sep 27;8(1):14491. doi: 10.1038/s41598-018-32820-5.
  PUBMED    30262876
REFERENCE   2  (bases 1 to 6425)
  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: "Sci Rep."; 
            date: "2018-09-27"; volume: "8(1):14491. doi"; pages: " 
            10.1038/s41598-018-32820-5"
COMMENT     Sequence Label: pCAG-Apex2-Venus-CAAX
FEATURES             Location/Qualifiers
     source          1..6425
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1678..1697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1728..1751
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1752..2498
                     /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
                     EVTGGPKVPFHPGREDKPEPPPEGRLPDPTKGSDHLRDVFGKAMGLTDQDIVALSGGHT
                     IGAAHKERSGFEGPWTSNPLIFDNSYFTELLSGEKEGLLQLPSDKALLSDPVFRPLVDK
                     YAADEDAFFADYAEAHQKLSELGFADA"
     CDS             2505..2531
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     regulatory      2547..2556
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2553..2555,2556..2558,2559..3269)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2598..2619)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2859..2878)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3206..3227
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3449..3468)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3514..3569
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3568..3587)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3928..3944)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3928..3944)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3941..3963)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3952..3968
                     /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(3976..3982,3983..4000,4001..4006))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4021..4042
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4101..4296
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4147..4282
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4302..4436
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4339..4358)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4393..4412
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4504..4521)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4675..5263)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4755..4774)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5434..6225,6226..6294))
                     /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     6057..6076
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6295..6399)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attcatggac tacaaggatg
     1741 acgacgataa gggaaagtct tacccaactg tgagtgctga ttaccaggac gccgttgaga
     1801 aggcgaagaa gaagctcaga ggcttcatcg ctgagaagag atgcgctcct ctaatgctcc
     1861 gtttggcatt ccactctgct ggaacctttg acaagggcac gaagaccggt ggacccttcg
     1921 gaaccatcaa gcaccctgcc gaactggctc acagcgctaa caacggtctt gacatcgctg
     1981 ttaggctttt ggagccactc aaggcggagt tccctatttt gagctacgcc gatttctacc
     2041 agttggctgg cgttgttgcc gttgaggtca cgggtggacc taaggttcca ttccaccctg
     2101 gaagagagga caagcctgag ccaccaccag agggtcgctt gcccgatccc actaagggtt
     2161 ctgaccattt gagagatgtg tttggcaaag ctatggggct tactgaccaa gatatcgttg
     2221 ctctatctgg gggtcacact attggagctg cacacaagga gcgttctgga tttgagggtc
     2281 cctggacctc taatcctctt attttcgaca actcatactt cacggagttg ttgagtggtg
     2341 agaaggaagg tctccttcag ctaccttctg acaaggctct tttgtctgac cctgtattcc
     2401 gccctctcgt tgacaaatat gcagcggacg aagatgcctt ctttgctgat tacgctgagg
     2461 ctcaccaaaa gctttccgag cttgggtttg ctgatgccct gcagctgcct cccctggagc
     2521 gcctgaccct ggacggaatt ccggtcgcca ccatggtgag caagggcgag gagctgttca
     2581 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     2641 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ctgatctgca
     2701 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgggc tacggcctgc
     2761 agtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     2821 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     2881 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     2941 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     3001 acgtctatat caccgccgac aagcagaaga acggcatcaa ggccaacttc aagatccgcc
     3061 acaacatcga ggacggcggc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     3121 gcgacggccc cgtgctgctg cccgacaacc actacctgag ctaccagtcc gccctgagca
     3181 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     3241 tcactctcgg catggacgag ctgtacaagt ccggactcag atctaagctg aaccctcctg
     3301 atgagagtgg ccccggctgc atgagctgca agtgtgtgct ctcctgagga tctcgaggat
     3361 atcgcggccg cactcctcag gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca
     3421 atgccctggc tcacaaatac cactgagatc tttttccctc tgccaaaaat tatggggaca
     3481 tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa
     3541 tagtgtgttg gaattttttg tgtctctcac tcggaaggac atatgggagg gcaaatcatt
     3601 taaaacatca gaatgagtat ttggtttaga gtttggcaac atatgccata tgctggctgc
     3661 catgaacaaa ggtggctata aagaggtcat cagtatatga aacagccccc tgctgtccat
     3721 tccttattcc atagaaaagc cttgacttga ggttagattt tttttatatt ttgttttgtg
     3781 ttattttttt ctttaacatc cctaaaattt tccttacatg ttttactagc cagatttttc
     3841 ctcctctcct gactactccc agtcatagct gtccctcttc tcttatgaag atccctcgac
     3901 ctgcagccca agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     3961 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     4021 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     4081 cctgtcgtgc cagcggatcc gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     4141 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     4201 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     4261 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc taacttgttt attgcagctt
     4321 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     4381 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatccgct
     4441 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     4501 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     4561 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     4621 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     4681 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     4741 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     4801 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     4861 gctttctcaa tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     4921 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     4981 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     5041 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5101 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5161 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     5221 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     5281 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     5341 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     5401 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     5461 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     5521 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     5581 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     5641 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     5701 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     5761 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     5821 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     5881 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     5941 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6001 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6061 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6121 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6181 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6241 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     6301 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     6361 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6421 acctg
//
