LOCUS       sequence-449277-        6232 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mamalian cell expression of cytosolic mEGFP-SNAP-tag2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6232)
  AUTHORS   Kuhn S, Nasufovic V, Wilhelm J, Kompa J, de Lange EMF, Lin YH, 
            Egoldt C, Fischer J, Lennoi A, Tarnawski M, Reinstein J, Vlijm R, 
            Hiblot J, Johnsson K
  TITLE     SNAP-tag2 for faster and brighter protein labeling.
  JOURNAL   Nat Chem Biol. 2025 Jul 3. doi: 10.1038/s41589-025-01942-z.
  PUBMED    40610720
REFERENCE   2  (bases 1 to 6232)
  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 Chem 
            Biol. 2025 Jul 3. doi: 10.1038/s41589-025-01942-z."
COMMENT     Sequence Label: pcDNA5/FRT_mEGFP-SNAP-tag2
FEATURES             Location/Qualifiers
     source          1..6232
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(66..82)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(66..82)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(79..101)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    90..106
                     /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(114..120,121..138,139..144))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    159..180
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(297..314)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(468..1056)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(548..567)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1227..2018,2019..2087))
                     /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     1850..1869
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2088..2192)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2267..2286)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2458..2837
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2838..3041
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2992..3012
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3086..3105
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3086..3104
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(3140..3142,3143..3145,3146..3856)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3185..3206)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3446..3465)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3793..3814
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4407..4424)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4413..4637
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4770..4789)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4921..4968
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4976..5998
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4976..5998
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    join(6128..6232,1..17)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6165..6184)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     join(6219..6232,1..6)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
       61 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
      121 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
      181 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
      241 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
      301 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
      361 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
      421 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
      481 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
      541 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
      601 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
      661 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
      721 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
      781 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
      841 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
      901 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
      961 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     1021 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     1081 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     1141 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     1201 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     1261 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     1321 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     1381 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     1441 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     1501 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     1561 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     1621 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     1681 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     1741 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     1801 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     1861 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     1921 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     1981 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     2041 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     2101 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     2161 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     2221 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
     2281 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
     2341 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
     2401 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
     2461 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
     2521 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
     2581 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
     2641 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
     2701 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
     2761 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
     2821 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
     2881 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
     2941 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
     3001 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tctctggcta actagagaac
     3061 ccactgctta ctggcttatc gaaattaata cgactcacta tagggagacc caagctggct
     3121 agcgtttaaa cttaccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     3181 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
     3241 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
     3301 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
     3361 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
     3421 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     3481 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     3541 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
     3601 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
     3661 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     3721 gctgctgccc gacaaccact acctgagcac ccagtccaaa ctgagcaaag accccaacga
     3781 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     3841 ggacgagctg tacaagatgg ataaagactg cgaaatgaaa cgcactacct atgacagccc
     3901 gctgggtaaa ctgctgctga gcggttgcga gcagggcctg caccgcatct atttcgtggg
     3961 caatggtcag ggcgagcagg gtcctccggg cccggagccg ctgatgcagg ctaccgcatg
     4021 gctgaacgca tattttcacc agccagaggc gattgaagag ttccctgtac cggccctgca
     4081 tcacccggta tttcagcagg aatcctttac ccgtcaggta ctgtggaaac tgctgaaagt
     4141 agtgaaattc ggcgaggtga tctcctattc ccagctggcg gcgctggctg gcaacccagc
     4201 ggcaactgcg gcggtgaaaa ccgctctgcg tggtaatccg gtgccaattc tgatcccgtg
     4261 ccaccgtgtg gttcagggcg acggcgacgt aggtggctac gaaggccctc tgtacgtgaa
     4321 agaatggctg ctggcgcacg aaggtcaccg tctgggtaaa ccaggcctgg gctagtctag
     4381 agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     4441 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     4501 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     4561 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     4621 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
     4681 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     4741 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     4801 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     4861 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtaccta
     4921 gaagttccta ttccgaagtt cctattctct agaaagtata ggaacttcct tggccaaaaa
     4981 gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc
     5041 cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg
     5101 gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt
     5161 ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt
     5221 cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct
     5281 gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc
     5341 tgcggccgat cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca
     5401 atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca
     5461 aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct
     5521 ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa
     5581 tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg
     5641 ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga
     5701 gcagcagacg cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg
     5761 ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt
     5821 cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac
     5881 tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga
     5941 agtactcgcc gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagca
     6001 cgtactacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt
     6061 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     6121 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     6181 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aa
//
