LOCUS       sequence-335908-        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        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      81..90
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             87..794
                     /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(232..250)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     500..519
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             813..1355
                     /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    2738..2872
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2762..2781)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2816..2835
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2985..3005)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2985..3003)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3024..3040)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3024..3040)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3037..3059)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3048..3064
                     /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(3072..3078,3079..3096,3097..3102))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3117..3138
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3255..3272)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3426..4014)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3506..3525)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4185..4976,4977..5045))
                     /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     4808..4827
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5046..5150)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5176..5631)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5313..5334)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5525..5544
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5702..6686
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagt gccaccatgg tgagcaaggg cgaggaggat aacatggcca
      121 tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac ggccacgagt
      181 tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga
      241 aggtgaccaa gggtggcccc ctgcccttcg cctgggacat cctgtcccct cagttcatgt
      301 acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg aagctgtcct
      361 tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg
      421 tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag ctgcgcggca
      481 ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg gaggcctcct
      541 ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagcag aggctgaagc
      601 tgaaggacgg cggccactac gacgctgagg tcaagaccac ctacaaggcc aagaagcccg
      661 tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga catcacctcc cacaacgagg
      721 actacaccat cgtggaacag tacgaacgcg ccgagggccg ccactccacc ggcggcatgg
      781 acgagctgta caagggtact ggctctggaa ccgacaaaga ctgcgaaatg aagcgcacca
      841 ccctggatag ccctctgggc aagctggaac tgtctgggtg cgaacagggc ctgcacgaga
      901 tcaagctgct gggcaaagga acatctgccg ccgacgccgt ggaagtgcct gccccagccg
      961 ccgtgctggg cggaccagag ccactgatgc aggccaccgc ctggctcaac gcctactttc
     1021 accagcctga ggccatcgag gagttccctg tgccagccct gcaccaccca gtgttccagc
     1081 aggagagctt tacccgccag gtgctgtgga aactgctgaa agtggtgaag ttcggagagg
     1141 tcatcagcta ccagcagctg gccgccctgg ccggcaatcc cgccgccacc gccgccgtga
     1201 aaaccgccct gagcggaaat cccgtgccca ttctgatccc cgcccaccgg gtggtgtcta
     1261 gctctggcgc cgtggggggc tacgagggcg ggctcgccgt gaaagagtgg ctgctggccc
     1321 acgagggcca cagactgggc aagcctgggc tgggtgggag tggaacaggc agcggtaccg
     1381 gctcagataa ggactgcgag atgaaacgga ccacactgga ctcaccactg gggaaactgg
     1441 agctgagcgg ctgcgagcag ggactccacg aaatcaaact gctcggcaag gggactagcg
     1501 ccgctgatgc tgtggaggtc ccagctcctg cagccgtcct gggaggaccc gaacccctca
     1561 tgcaagctac cgcatggctg aacgcatact tccatcagcc agaagctatc gaagaatttc
     1621 ccgtgcccgc cctccaccat cccgtctttc agcaagaaag tttcacacgg caggtcctct
     1681 ggaagctcct gaaggtggtc aaattcggcg aagtgatctc atatcagcaa ctcgctgcac
     1741 tggcaggaaa ccctgcagct accgcagcag tgaagacagc tctgtctggg aaccccgtcc
     1801 ctatcctgat tcctgcccat agagtggtca gctccagcgg agcagtggga ggatatgagg
     1861 gaggactggc tgtcaaggaa tggctcctgg cacatgaagg ccataggctg ggaaaacccg
     1921 gactgggacc cgccgggatt ggagcacctg ggagcctcga gacgggttct gggacaggat
     1981 cgggctctgc ttggaagcca gacgaaacgg ctccacagag ttccctgagg cgcccagacc
     2041 ccggccggct tccaccagct gccagcgagg aagcacacac cagcaatgtc aagatggaaa
     2101 aattgtacct gcattacttg agagcagaga gctttagaaa agctctgatt tatcaaaaga
     2161 agtatctttt gctgttgatt ggtggattcc aggattctga acaagaaaca ctctccatga
     2221 ttgcccattt gggggtattt ccttccaaag cagaacggaa aatcacatct cgtcctttca
     2281 ccaggttccg cacggccgtc agggtggtca ttgcaatatt aagattacgt tttttggtta
     2341 agaaatggca agaagtagat cggaaaggag ctctggcaca aggcaaagcc cctcgcccag
     2401 ggccccgagc acgacagccg cagtctccac ccagaaccag agagtccccc ccaacccggg
     2461 atgtaccctc tggccacacc agggaccctg ccagaggccg cagactggca gcagcagcct
     2521 ccccacacag tgggggaaga gccactccat ccccaaattc aagattagaa agatccctga
     2581 ctgcttctca agatccagaa cattccttga cagagtatat tcaccattta gaagtgatcc
     2641 agcaaagatt gggaggggta ctaccagatt ctacttcaaa gaaatcctgc cacccgatga
     2701 acctgcagtc tagaactata gtgagtcgta ttacgtagat ccagacatga taagatacat
     2761 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     2821 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa
     2881 caattgcatt cattttatgt ttcaggttca gggggaggtg tgggaggttt tttaattcgc
     2941 ggccgcggcg ccaatgcatt gggcccggta cccagctttt gttcccttta gtgagggtta
     3001 attgcgcgct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     3061 acaattccac acaacatacg agccgggagc ataaagtgta aagcctgggg tgcctaatga
     3121 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     3181 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     3241 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     3301 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     3361 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     3421 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     3481 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3541 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3601 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3661 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     3721 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     3781 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     3841 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3901 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     3961 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     4021 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     4081 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     4141 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     4201 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     4261 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     4321 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     4381 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     4441 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     4501 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     4561 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     4621 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     4681 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     4741 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     4801 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     4861 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     4921 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     4981 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     5041 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     5101 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     5161 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     5221 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     5281 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     5341 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     5401 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     5461 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     5521 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     5581 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc
     5641 cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc
     5701 agtcgaccgc caattcaata tggcgtatat ggactcatgc caattcaata tggtggatct
     5761 ggacctgtgc caattcaata tggcgtatat ggactcgtgc caattcaata tggtggatct
     5821 ggaccccagc caattcaata tggcggactt ggcaccatgc caattcaata tggcggacct
     5881 ggcactgtgc caactgggga ggggtctact tggcacggtg ccaagtttga ggaggggtct
     5941 tggccctgtg ccaagtccgc catattgaat tggcatggtg ccaataatgg cggccatatt
     6001 ggctatatgc caggatcaat atataggcaa tatccaatat ggccctatgc caatatggct
     6061 attggccagg ttcaatacta tgtattggcc ctatgccata tagtattcca tatatgggtt
     6121 ttcctattga cgtagatagc ccctcccaat gggcggtccc atataccata tatggggctt
     6181 cctaataccg cccatagcca ctcccccatt gacgtcaatg gtctctatat atggtctttc
     6241 ctattgacgt catatgggcg gtcctattga cgtatatggc gcctccccca ttgacgtcaa
     6301 ttacggtaaa tggcccgcct ggctcaatgc ccattgacgt caataggacc acccaccatt
     6361 gacgtcaatg ggatggctca ttgcccattc atatccgttc tcacgccccc tattgacgtc
     6421 aatgacggta aatggcccac ttggcagtac atcaatatct attaatagta acttggcaag
     6481 tacattacta ttggaagtac gccagggtac attggcagta ctcccattga cgtcaatggc
     6541 ggtaaatggc ccgcgatggc tgccaagtac atccccattg acgtcaatgg ggaggggcaa
     6601 tgacgcaaat gggcgttcca ttgacgtaaa tgggcggtag gcgtgcctaa tgggaggtct
     6661 atataagcaa tgctcgttta gggaac
//
