LOCUS       sequence-96168-e        6616 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses membrane-targeted SNAP(f)-tag and histone-GFP, 
            non-conditional.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6616)
  AUTHORS   Kohl J, Ng J, Cachero S, Ciabatti E, Dolan MJ, Sutcliffe B, Tozer A,
            Ruehle S, Krueger D, Frechter S, Branco T, Tripodi M, Jefferis GS
  TITLE     Ultrafast tissue staining with chemical tags.
  JOURNAL   Proc Natl Acad Sci U S A. 2014 Aug 25. pii: 201411087.
  PUBMED    25157152
REFERENCE   2  (bases 1 to 6616)
  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: "Proc Natl 
            Acad Sci U S A. 2014 Aug 25. pii: 201411087."
COMMENT     Sequence Label: pAAV-myrSNAP
FEATURES             Location/Qualifiers
     source          1..6616
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     enhancer        221..524
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        525..727
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     678..698
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     724..748
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          876..1254
                     /label=chimeric intron
                     /note="fusion between human cytomegalovirus intron A and 
                     human beta-globin intron 2"
     regulatory      1385..1394
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1454..1999
                     /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             2009..2062
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2081..2458
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAVTKYTSAK"
     CDS             join(2543..2545,2546..3256)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(2585..2606)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2846..2865)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3193..3214
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3363..3839
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3562..3581)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3879..4019
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3879..4008
                     /label=AAV2 ITR
     rep_origin      4094..4549
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4181..4200)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4391..4412
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4566..4585)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4685..4707
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4745..4763)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4831..4935
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4936..5004,5005..5796)
                     /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(5154..5173)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5967..6555
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6456..6475
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtggagc tagttattaa tagtaatcaa
      181 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
      241 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
      301 ttcccatagt aacgtcaata gggactttcc attgacgtca atgggtggag tatttacggt
      361 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
      421 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
      481 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
      541 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
      601 ttgacgtcaa tgggagtttg ttttgcacca aaatcaacgg gactttccaa aatgtcgtaa
      661 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
      721 cagagctcgt ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct
      781 ccatagaaga caccgggacc gatccagcct ccgcggattc gaatcccggc cgggaacggt
      841 gcattggaac gcggattccc cgtgccaaga gtgacgtaag taccgcctat agagtctata
      901 ggcccacaaa aaatgctttc ttcttttaat atactttttt gtttatctta tttctaatac
      961 tttccctaat ctctttcttt cagggcaata atgatacaat gtatcatgcc tctttgcacc
     1021 attctaaaga ataacagtga taatttctgg gttaaggcaa tagcaatatt tctgcatata
     1081 aatatttctg catataaatt gtaactgatg taagaggttt catattgcta atagcagcta
     1141 caatccagct accattctgc ttttatttta tggttgggat aaggctggat tattctgagt
     1201 ccaagctagg cccttttgct aatcatgttc atacctctta tcttcctccc acagctcctg
     1261 ggcaacgtgc tggtctgtgt gctggcccat cactttggca aagaattggg attcgaacat
     1321 cgattgaatt cccaccatgg gctgcatcaa gagcaagcgc aaggacaacc tgaacgacga
     1381 cgaggccgcc atgggctgca tcaagagcaa gcgcaaggac aacctgaacg acgacgaggc
     1441 accggtcgcc tccatggaca aagactgcga aatgaagcgc accaccctgg atagccctct
     1501 gggcaagctg gaactgtctg ggtgcgaaca gggcctgcac cgtatcatct tcctgggcaa
     1561 aggaacatct gccgccgacg ccgtggaagt gcctgcccca gccgccgtgc tgggcggacc
     1621 agagccactg atgcaggcca ccgcctggct caacgcctac tttcaccagc ctgaggccat
     1681 cgaggagttc cctgtgccag ccctgcacca cccagtgttc cagcaggaga gctttacccg
     1741 ccaggtgctg tggaaactgc tgaaagtggt gaagttcgga gaggtcatca gctacagcca
     1801 cctggccgcc ctggccggca atcccgccgc caccgccgcc gtgaaaaccg ccctgagcgg
     1861 aaatcccgtg cccattctga tcccctgcca ccgggtggtg cagggcgacc tggacgtggg
     1921 gggctacgag ggcgggctcg ccgtgaaaga gtggctgctg gcccacgagg gccacagact
     1981 gggcaagcct gggctgggtg gcagtggaga gggcagagga agtctgctaa catgcggtga
     2041 cgtcgaggag aatcctggcc caggagatct gaatatcacc atgccagagc cagcgaagtc
     2101 tgctcccgcc ccgaaaaagg gctccaagaa ggcggtgact aaggcgcaga agaaaggcgg
     2161 caagaagcgc aagcgcagcc gcaaggagag ctattccatc tatgtgtaca aggttctgaa
     2221 gcaggtccac cctgacaccg gcatttcgtc caaggccatg ggcatcatga actcgtttgt
     2281 gaacgacatt ttcgagcgca tcgcaggtga ggcttcccgc ctggcgcatt acaacaagcg
     2341 ctcgaccatc acctccaggg agatccagac ggccgtgcgc ctgctgctgc ctggggagtt
     2401 ggccaagcac gccgtgtccg agggtactaa ggccgtcacc aagtacacca gcgctaaggg
     2461 atctgggggt ggaggctccg gcgggggtgg atctggaggt gggggcagcg gcggaggtgg
     2521 gctagcgcta ccgcgggcca ccatgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     2581 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
     2641 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
     2701 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
     2761 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
     2821 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     2881 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     2941 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
     3001 ggccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga
     3061 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     3121 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
     3181 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctcacggcat
     3241 ggacgagctg tacaagtccg gactcagatc atcaagctca tcggatccgg gcggccgtga
     3301 taataggtga tagtaagtaa tagtgaaagc ttgcctcgag cagcgctgct cgagagatct
     3361 acgggtggca tccctgtgac ccctccccag tgcctctcct ggccctggaa gttgccactc
     3421 cagtgcccac cagccttgtc ctaataaaat taagttgcat cattttgtct gactaggtgt
     3481 ccttctataa tattatgggg tggagggggg tggtatggag caaggggcaa gttgggaaga
     3541 caacctgtag ggcctgcggg gtctattggg aaccaagctg gagtgcagtg gcacaatctt
     3601 ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt
     3661 tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac
     3721 ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg atctacccac
     3781 cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc
     3841 tgattttgta ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt
     3901 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     3961 acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg
     4021 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca
     4081 aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     4141 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     4201 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     4261 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt
     4321 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     4381 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcgggctat
     4441 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     4501 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact
     4561 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     4621 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     4681 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     4741 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     4801 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     4861 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     4921 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     4981 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5041 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5101 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     5161 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     5221 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     5281 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     5341 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     5401 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     5461 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     5521 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     5581 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     5641 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     5701 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     5761 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     5821 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     5881 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     5941 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     6001 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     6061 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     6121 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     6181 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     6241 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     6301 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     6361 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     6421 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     6481 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     6541 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     6601 ccttttgctc acatgt
//
