LOCUS       sequence-392532-        5243 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing fluorescent SNAPf-mTurquoise2 fusion protein in mammalian
            cells with localization in the whole cell..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5243)
  AUTHORS   Martin A, Rivera-Fuentes P
  TITLE     A general strategy to develop fluorogenic polymethine dyes for 
            bioimaging.
  JOURNAL   Nat Chem. 2024 Jan;16(1):28-35. doi: 10.1038/s41557-023-01367-y. 
            Epub 2023 Nov 27.
  PUBMED    38012391
REFERENCE   2  (bases 1 to 5243)
  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"; doi: 
            "10.1038/s41557-023-01367-y"; journalName: "Nat Chem"; date: 
            "2024-01"; volume: "16"; issue: "1"; pages: "28-35"
COMMENT     Sequence Label: SNAPf-mTurquoise2
FEATURES             Location/Qualifiers
     source          1..5243
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             610..1155
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     CDS             join(1192..1194,1195..1908)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)"
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1234..1255)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1495..1514)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1842..1863
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2031..2152
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2068..2087)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2122..2141
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2159..2614)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2296..2317)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2508..2527
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2641..2745
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2747..3104
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2747..2938
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2770..2790)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2955..3090
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3139..3933
                     /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
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFGHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3193..3212)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3803..3822
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4121..4140)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4165..4212
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4541..5129
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5030..5049
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcaa tggacaaaga ctgcgaaatg aagcgcacca ccctggatag ccctctgggc
      661 aagctggaac tgtctgggtg cgaacagggc ctgcaccgta tcatcttcct gggcaaagga
      721 acatctgccg ccgacgccgt ggaagtgcct gccccagccg ccgtgctggg cggaccagag
      781 ccactgatgc aggccaccgc ctggctcaac gcctactttc accagcctga ggccatcgag
      841 gagttccctg tgccagccct gcaccaccca gtgttccagc aggagagctt tacccgccag
      901 gtgctgtgga aactgctgaa agtggtgaag ttcggagagg tcatcagcta cagccacctg
      961 gccgccctgg ccggcaatcc cgccgccacc gccgccgtga aaaccgccct gagcggaaat
     1021 cccgtgccca ttctgatccc ctgccaccgg gtggtgcagg gcgacctgga cgtggggggc
     1081 tacgagggcg ggctcgccgt gaaagagtgg ctgctggccc acgagggcca cagactgggc
     1141 aagcctgggc tgggtcctgc aggcggatcc gcgtttaaac tcgaggttaa tgtgagcaag
     1201 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     1261 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     1321 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     1381 ctgtcctggg gcgtgcagtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
     1441 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     1501 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     1561 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     1621 aactacttta gcgacaacgt ctatatcacc gccgacaagc agaagaacgg catcaaggcc
     1681 aacttcaaga tccgccacaa catcgaggac ggcggcgtgc agctcgccga ccactaccag
     1741 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     1801 cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     1861 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaaag cggccgcgac
     1921 tctagatcat aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc
     1981 cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta
     2041 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     2101 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct taaggcgtaa
     2161 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
     2221 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
     2281 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
     2341 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
     2401 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     2461 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     2521 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     2581 cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg
     2641 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     2701 caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga
     2761 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     2821 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     2881 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     2941 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3001 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     3061 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca
     3121 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     3181 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     3241 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     3301 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
     3361 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     3421 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     3481 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcggc
     3541 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     3601 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     3661 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     3721 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     3781 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     3841 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     3901 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     3961 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     4021 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     4081 tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa
     4141 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg
     4201 ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac
     4261 cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa
     4321 gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct
     4381 caggttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4441 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4501 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     4561 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4621 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4681 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4741 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4801 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4861 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4921 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4981 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     5041 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     5101 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     5161 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     5221 ataaccgtat taccgccatg cat
//
