LOCUS       sequence-361820-        7534 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent expression of H2B-APEX fusion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7534)
  AUTHORS   Dumrongprechachan V, Salisbury RB, Soto G, Kumar M, MacDonald ML, 
            Kozorovitskiy Y
  TITLE     Cell-type and subcellular compartment-specific APEX2 proximity 
            labeling reveals activity-dependent nuclear proteome dynamics in the
            striatum.
  JOURNAL   Nat Commun. 2021 Aug 11;12(1):4855. doi: 10.1038/s41467-021-25144-y.
  PUBMED    34381044
REFERENCE   2  (bases 1 to 7534)
  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 
            Commun."; date: "2021-08-11"; volume: "12(1):4855. doi"; pages: " 
            10.1038/s41467-021-25144-y"
COMMENT     Sequence Label: pAAV-EF1a-DIO-H2B.APEX-P2A-EGFP
FEATURES             Location/Qualifiers
     source          1..7534
                     /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(join(1575..2285,2286..2288,2289..2291))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1575..2285,2286..2288,2289..2291))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1617..1638)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1966..1985
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2225..2246
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(2292..2348)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(2358..3104)
                     /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             complement(3105..3128)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(3129..3506)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     protein_bind    3519..3552
                     /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    3603..3636
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3661..4249
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3714..3734)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4132..4143)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4281..4757
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4480..4499)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4797..4937
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4797..4926
                     /label=AAV2 ITR
     rep_origin      5012..5467
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5099..5118)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5309..5330
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5484..5503)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5603..5625
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5663..5681)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5749..5853
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5854..5922,5923..6714)
                     /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(6072..6091)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6885..7473
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7374..7393
                     /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 gcgccttaac cggtcttgta cagctcgtcc atgccgagag tgatcccggc ggcggtcacg
     1621 aactccagca ggaccatgtg atcgcgcttc tcgttggggt ctttgctcag ggcggactgg
     1681 gtgctcaggt agtggttgtc gggcagcagc acggggccgt cgccgatggg ggtgttctgc
     1741 tggtagtggt cggcgagctg cacgctgccg tcctcgatgt tgtggcggat cttgaagttc
     1801 accttgatgc cgttcttctg cttgtcggcc atgatataga cgttgtggct gttgtagttg
     1861 tactccagct tgtgccccag gatgttgccg tcctccttga agtcgatgcc cttcagctcg
     1921 atgcggttca ccagggtgtc gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg
     1981 tcgtccttga agaagatggt gcgctcctgg acgtagcctt cgggcatggc ggacttgaag
     2041 aagtcgtgct gcttcatgtg gtcggggtag cggctgaagc actgcacgcc gtaggtcagg
     2101 gtggtcacga gggtgggcca gggcacgggc agcttgccgg tggtgcagat gaacttcagg
     2161 gtcagcttgc cgtaggtggc atcgccctcg ccctcgccgg acacgctgaa cttgtggccg
     2221 tttacgtcgc cgtccagctc gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc
     2281 ttgctcacca taggaccggg gttttcttcc acgtctcctg cttgctttaa cagagagaag
     2341 ttcgtggctc cggatccggc atcagcaaac ccaagctcgg aaagcttttg gtgagcctca
     2401 gcgtaatcag caaagaaggc atcttcgtcc gctgcatatt tgtcaacgag agggcggaat
     2461 acagggtcag acaaaagagc cttgtcagaa ggtagctgaa ggagaccttc cttctcacca
     2521 ctcaacaact ccgtgaagta tgagttgtcg aaaataagag gattagaggt ccagggaccc
     2581 tcaaatccag aacgctcctt gtgtgcagct ccaatagtgt gacccccaga tagagcaacg
     2641 atatcttggt cagtaagccc catagctttg ccaaacacat ctctcaaatg gtcagaaccc
     2701 ttagtgggat cgggcaagcg accctctggt ggtggctcag gcttgtcctc tcttccaggg
     2761 tggaatggaa ccttaggtcc acccgtgacc tcaacggcaa caacgccagc caactggtag
     2821 aaatcggcgt agctcaaaat agggaactcc gccttgagtg gctccaaaag cctaacagcg
     2881 atgtcaagac cgttgttagc gctgtgagcc agttcggcag ggtgcttgat ggttccgaag
     2941 ggtccaccgg tcttcgtgcc cttgtcaaag gttccagcag agtggaatgc caaacggagc
     3001 attagaggag cgcatctctt ctcagcgatg aagcctctga gcttcttctt cgccttctca
     3061 acggcgtcct ggtaatcagc actcacagtt gggtaagact ttcccttatc gtcgtcatcc
     3121 ttgtagtcct tagcgctggt gtacttggtg atggccttag taccctcgga cacggcgtgc
     3181 ttggccaact ccccaggcag cagcaggcgc acggccgtct ggatctccct ggaggtgatg
     3241 gtcgagcgct tgttgtaatg cgccaggcgg gaagcctcac ctgcgatgcg ctcgaaaatg
     3301 tcgttcacaa acgaattcat gatgcccatg gccttggacg aaatgccggt gtcagggtgg
     3361 acctgcttca gaaccttgta cacatagatg gaatagctct ccttgcggct gcgcttgcgc
     3421 ttcttgccgc ctttcttctg cgccttagtc accgccttct tggagccctt tttcggggcg
     3481 ggagcagact tcgctggctc tggcatggtg gcgctagcat aacttcgtat aaagtatcct
     3541 atacgaagtt atttgcctta acccagaaat tatcactgtt attctttaga atggtgcaaa
     3601 gaataacttc gtataatgta tgctatacga agttatgaat tcgatatcaa gcttatcgat
     3661 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     3721 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     3781 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     3841 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     3901 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     3961 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     4021 ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc
     4081 gcctatgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     4141 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     4201 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccga
     4261 gcgctgctcg agagatctac gggtggcatc cctgtgaccc ctccccagtg cctctcctgg
     4321 ccctggaagt tgccactcca gtgcccacca gccttgtcct aataaaatta agttgcatca
     4381 ttttgtctga ctaggtgtcc ttctataata ttatggggtg gaggggggtg gtatggagca
     4441 aggggcaagt tgggaagaca acctgtaggg cctgcggggt ctattgggaa ccaagctgga
     4501 gtgcagtggc acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc
     4561 tgcctcagcc tcccgagttg ttgggattcc aggcatgcat gaccaggctc agctaatttt
     4621 tgtttttttg gtagagacgg ggtttcacca tattggccag gctggtctcc aactcctaat
     4681 ctcaggtgat ctacccacct tggcctccca aattgctggg attacaggcg tgaaccactg
     4741 ctcccttccc tgtccttctg attttgtagg taaccacgtg cggaccgagc ggccgcagga
     4801 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
     4861 gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc
     4921 gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     4981 ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg
     5041 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgcctt agcgcccgct
     5101 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     5161 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     5221 cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     5281 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     5341 aactctatct cgggctattc ttttgattta taagggattt tgccgatttc ggtctattgg
     5401 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt
     5461 acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     5521 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     5581 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     5641 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     5701 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     5761 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     5821 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     5881 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     5941 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     6001 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     6061 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     6121 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     6181 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     6241 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     6301 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     6361 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     6421 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     6481 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     6541 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     6601 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     6661 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     6721 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     6781 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     6841 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     6901 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     6961 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     7021 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     7081 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     7141 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     7201 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     7261 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     7321 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     7381 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     7441 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     7501 ttcctggcct tttgctggcc ttttgctcac atgt
//
