LOCUS       sequence-257388-        4696 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Localization: N1 Cloning Vector, Excitation: 505 / 569, Emission: 
            516 / 581.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4696)
  AUTHORS   Paez-Segala MG, Sun MG, Shtengel G, Viswanathan S, Baird MA, Macklin
            JJ, Patel R, Allen JR, Howe ES, Piszczek G, Hess HF, Davidson MW, 
            Wang Y, Looger LL
  TITLE     Fixation-resistant photoactivatable fluorescent proteins for CLEM.
  JOURNAL   Nat Methods. 2015 Jan 12. doi: 10.1038/nmeth.3225.
  PUBMED    25581799
REFERENCE   2  (bases 1 to 4696)
  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 
            Methods. 2015 Jan 12. doi: 10.1038/nmeth.3225."
COMMENT     Sequence Label: mEos4a-N1
FEATURES             Location/Qualifiers
     source          1..4696
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        85..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        389..592
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     543..563
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    615..695
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      697..706
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             703..1386
                     /codon_start=1
                     /product="fixation-resistant green-to-red photoswitchable 
                     fluorescent protein (Paez-Segala et al., 2015)"
                     /label=mEos4a
                     /note="derived from mEos2"
                     /translation="MVSAIKPDMRIKLRMEGNVNGHHFVIDGDGTGKPYEGKQTMDLEV
                     KEGGPLPFAFDILTTAFHYGNRVFVKYPDNIQDYFKQSFPKGYSWERSLTFEDGGICYA
                     RNDITMEGDTFYNKVRFYGTNFPANGPVMQKKTLKWEPSTEKMYVRDGVLTGDIHMALL
                     LEGNAHYRCDFRTTYKAKEKGVKLPGYHFVDHAIEILSHDKDYNKVKLYEHAVAHSGLP
                     DNARR"
     polyA_signal    1508..1629
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1545..1564)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1599..1618
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1636..2091)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1773..1794)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1985..2004
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2118..2222
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2224..2581
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2224..2415
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2247..2267)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2432..2567
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2616..3410
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2616..3410
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2670..2689)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3280..3299
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3598..3617)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3642..3689
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4018..4606
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4507..4526
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gataaccgta ttaccgccat gcattagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      421 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      481 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      541 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt
      601 gaaccgtcag atccgctagc gctaccggac tcagatctcg agctcaagct tcgaattctg
      661 cagtcgacgg taccgcgggc ccgggatcca ccggtcgcca ccatggtgag tgcgattaag
      721 ccagacatga ggatcaaact ccgtatggaa ggcaacgtaa acgggcacca ctttgtgatc
      781 gacggagatg gtacaggcaa gccttatgag ggaaaacaga ccatggatct tgaagtcaaa
      841 gagggcggac ctctgccttt tgcctttgat atcctgacca ctgcattcca ttacggcaac
      901 agggtattcg ttaaatatcc agacaacata caagactatt ttaagcagtc gtttcctaag
      961 gggtattcgt gggaacgaag cttgactttc gaagacgggg gcatttgcta tgccagaaac
     1021 gacataacaa tggaagggga cactttctat aataaagttc gattttatgg taccaacttt
     1081 cccgccaatg gtccagttat gcagaagaag acgctgaaat gggagccctc cactgagaaa
     1141 atgtatgtgc gtgatggagt gctgacgggt gatattcata tggctttgtt gcttgaagga
     1201 aatgcccatt accgatgtga cttcagaact acttacaaag ctaaggagaa gggtgtcaag
     1261 ttaccaggct accactttgt ggaccacgcc attgagattt taagccatga caaagattac
     1321 aacaaggtta agctgtatga gcatgctgtt gctcattctg gattgcctga caatgccaga
     1381 cgataagcgg ccgcgactct agatcataat cagccatacc acatttgtag aggttttact
     1441 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     1501 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     1561 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     1621 tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     1681 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     1741 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     1801 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     1861 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     1921 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     1981 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     2041 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     2101 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     2161 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     2221 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     2281 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     2341 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2401 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     2461 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     2521 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     2581 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     2641 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     2701 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     2761 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     2821 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     2881 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     2941 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     3001 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     3061 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     3121 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     3181 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     3241 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     3301 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     3361 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     3421 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     3481 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     3541 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     3601 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     3661 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     3721 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     3781 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     3841 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     3901 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     3961 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     4021 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     4081 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     4141 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     4201 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     4261 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     4321 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     4381 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     4441 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     4501 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     4561 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     4621 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     4681 atcccctgat tctgtg
//
