LOCUS       sequence-339784-        6686 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dead SNAP tag at the centrosome.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6686)
  AUTHORS   Bucko PJ, Lombard CK, Rathbun L, Garcia I, Bhat A, Wordeman L, Smith
            FD, Maly DJ, Hehnly H, Scott JD
  TITLE     Subcellular drug targeting illuminates local kinase action.
  JOURNAL   Elife. 2019 Dec 24;8. pii: 52220. doi: 10.7554/eLife.52220.
  PUBMED    31872801
REFERENCE   2  (bases 1 to 6686)
  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: "Elife."; 
            date: "2019-12-24"; volume: "8. pii: 52220. doi"; pages: " 
            10.7554/eLife.52220"
COMMENT     Sequence Label: pCS2-mCherry_SNAP_SNAPV_PACT_Dead
FEATURES             Location/Qualifiers
     source          1..6686
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..978
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1013..1031
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1013..1030
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      1059..1068
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1065..1772
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1210..1228)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1478..1497
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1791..2333
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="optimized for mammalian expression"
                     /translation="DKDCEMKRTTLDSPLGKLELSGCEQGLHEIKLLGKGTSAADAVEV
                     PAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLKV
                     VKFGEVISYQQLAALAGNPAATAAVKTALSGNPVPILIPAHRVVSSSGAVGGYEGGLAV
                     KEWLLAHEGHRLGKPGLG"
     polyA_signal    3716..3850
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3740..3759)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3794..3813
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3963..3983)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3963..3981)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4002..4018)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4002..4018)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4015..4037)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4026..4042
                     /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(4050..4056,4057..4074,4075..4080))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4095..4116
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4233..4250)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4404..4992)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4484..4503)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5163..5954,5955..6023))
                     /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     5786..5805
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6024..6128)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6154..6609)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6291..6312)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6503..6522
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gccaattcaa tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt
       61 gccaattcaa tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca
      121 gccaattcaa tatggcggac ttggcaccat gccaattcaa tatggcggac ctggcactgt
      181 gccaactggg gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg
      241 tgccaagtcc gccatattga attggcatgg tgccaataat ggcggccata ttggctatat
      301 gccaggatca atatataggc aatatccaat atggccctat gccaatatgg ctattggcca
      361 ggttcaatac tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt
      421 gacgtagata gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac
      481 cgcccatagc cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac
      541 gtcatatggg cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta
      601 aatggcccgc ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa
      661 tgggatggct cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg
      721 taaatggccc acttggcagt acatcaatat ctattaatag taacttggca agtacattac
      781 tattggaagt acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg
      841 gcccgcgatg gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa
      901 atgggcgttc cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc
      961 aatgctcgtt tagggaaccg ccattctgcc tggggacgtc ggagcaagct tgatttaggt
     1021 gacactatag aatacaagct acttgttctt tttgcagtgc caccatggtg agcaagggcg
     1081 aggaggataa catggccatc atcaaggagt tcatgcgctt caaggtgcac atggagggct
     1141 ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
     1201 cccagaccgc caagctgaag gtgaccaagg gtggccccct gcccttcgcc tgggacatcc
     1261 tgtcccctca gttcatgtac ggctccaagg cctacgtgaa gcaccccgcc gacatccccg
     1321 actacttgaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg
     1381 acggcggcgt ggtgaccgtg acccaggact cctccctgca ggacggcgag ttcatctaca
     1441 aggtgaagct gcgcggcacc aacttcccct ccgacggccc cgtaatgcag aagaagacca
     1501 tgggctggga ggcctcctcc gagcggatgt accccgagga cggcgccctg aagggcgaga
     1561 tcaagcagag gctgaagctg aaggacggcg gccactacga cgctgaggtc aagaccacct
     1621 acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa cgtcaacatc aagttggaca
     1681 tcacctccca caacgaggac tacaccatcg tggaacagta cgaacgcgcc gagggccgcc
     1741 actccaccgg cggcatggac gagctgtaca agggtactgg ctctggaacc gacaaagact
     1801 gcgaaatgaa gcgcaccacc ctggatagcc ctctgggcaa gctggaactg tctgggtgcg
     1861 aacagggcct gcacgagatc aagctgctgg gcaaaggaac atctgccgcc gacgccgtgg
     1921 aagtgcctgc cccagccgcc gtgctgggcg gaccagagcc actgatgcag gccaccgcct
     1981 ggctcaacgc ctactttcac cagcctgagg ccatcgagga gttccctgtg ccagccctgc
     2041 accacccagt gttccagcag gagagcttta cccgccaggt gctgtggaaa ctgctgaaag
     2101 tggtgaagtt cggagaggtc atcagctacc agcagctggc cgccctggcc ggcaatcccg
     2161 ccgccaccgc cgccgtgaaa accgccctga gcggaaatcc cgtgcccatt ctgatccccg
     2221 cccaccgggt ggtgtctagc tctggcgccg tggggggcta cgagggcggg ctcgccgtga
     2281 aagagtggct gctggcccac gagggccaca gactgggcaa gcctgggctg ggtgggagtg
     2341 gaacaggcag cggtaccggc tcagataagg actgcgagat gaaacggacc acactggact
     2401 caccactggg gaaactggag ctgagcggct gcgagcaggg actccacgaa atcaaactgc
     2461 tcggcaaggg gactagcgcc gctgatgctg tggaggtccc agctcctgca gccgtcctgg
     2521 gaggacccga acccctcatg caagctaccg catggctgaa cgcatacttc catcagccag
     2581 aagctatcga agaatttccc gtgcccgccc tccaccatcc cgtctttcag caagaaagtt
     2641 tcacacggca ggtcctctgg aagctcctga aggtggtcaa attcggcgaa gtgatctcat
     2701 atcagcaact cgctgcactg gcaggaaacc ctgcagctac cgcagcagtg aagacagctc
     2761 tgtctgggaa ccccgtccct atcctgattc ctgcccatag agtggtcagc tccagcggag
     2821 cagtgggagg atatgaggga ggactggctg tcaaggaatg gctcctggca catgaaggcc
     2881 ataggctggg aaaacccgga ctgggacccg ccgggattgg agcacctggg agcctcgaga
     2941 cgggttctgg gacaggatcg ggctctgctt ggaagccaga cgaaacggct ccacagagtt
     3001 ccctgaggcg cccagacccc ggccggcttc caccagctgc cagcgaggaa gcacacacca
     3061 gcaatgtcaa gatggaaaaa ttgtacctgc attacttgag agcagagagc tttagaaaag
     3121 ctctgattta tcaaaagaag tatcttttgc tgttgattgg tggattccag gattctgaac
     3181 aagaaacact ctccatgatt gcccatttgg gggtatttcc ttccaaagca gaacggaaaa
     3241 tcacatctcg tcctttcacc aggttccgca cggccgtcag ggtggtcatt gcaatattaa
     3301 gattacgttt tttggttaag aaatggcaag aagtagatcg gaaaggagct ctggcacaag
     3361 gcaaagcccc tcgcccaggg ccccgagcac gacagccgca gtctccaccc agaaccagag
     3421 agtccccccc aacccgggat gtaccctctg gccacaccag ggaccctgcc agaggccgca
     3481 gactggcagc agcagcctcc ccacacagtg ggggaagagc cactccatcc ccaaattcaa
     3541 gattagaaag atccctgact gcttctcaag atccagaaca ttccttgaca gagtatattc
     3601 accatttaga agtgatccag caaagattgg gaggggtact accagattct acttcaaaga
     3661 aatcctgcca cccgatgaac ctgcagtcta gaactatagt gagtcgtatt acgtagatcc
     3721 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     3781 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     3841 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     3901 ggaggttttt taattcgcgg ccgcggcgcc aatgcattgg gcccggtacc cagcttttgt
     3961 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     4021 tgaaattgtt atccgctcac aattccacac aacatacgag ccgggagcat aaagtgtaaa
     4081 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     4141 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     4201 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     4261 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     4321 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     4381 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     4441 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     4501 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     4561 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     4621 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     4681 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     4741 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     4801 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     4861 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     4921 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     4981 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     5041 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     5101 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     5161 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     5221 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     5281 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     5341 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     5401 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     5461 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     5521 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     5581 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     5641 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     5701 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     5761 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     5821 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     5881 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     5941 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     6001 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     6061 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6121 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     6181 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     6241 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     6301 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     6361 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     6421 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     6481 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     6541 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     6601 agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     6661 cctcttcgct attacgccag tcgacc
//
