LOCUS       sequence-451905-        6455 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For transient expression of the tdMCP-protein standard. Used in 
            LABEL-seq abundance assays..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6455)
  AUTHORS   Simon JJ, Fowler DM, Maly DJ
  TITLE     Multiplexed profiling of intracellular protein abundance, activity, 
            interactions and druggability with LABEL-seq.
  JOURNAL   Nat Methods. 2024 Nov;21(11):2094-2106. doi: 
            10.1038/s41592-024-02456-7. Epub 2024 Oct 21.
  PUBMED    39433876
REFERENCE   2  (bases 1 to 6455)
  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/s41592-024-02456-7"; journalName: "Nat Methods"; date: 
            "2024-11"; volume: "21"; issue: "11"; pages: "2094-2106"
COMMENT     Sequence Label: 2xMyc-tdMCP-SNAP (tdMCP standard)
FEATURES             Location/Qualifiers
     source          1..6455
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      234..822
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     723..742
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     976..993
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1110..1131
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1146..1151,1152..1169,1170..1176)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1184..1200
                     /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)."
     primer_bind     1189..1211
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1208..1224
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1208..1224
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1273..1394
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1284..1303)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1338..1357
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1524..2546)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             complement(1524..2546)
                     /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
                     "
     protein_bind    complement(2554..2601)
                     /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)."
     primer_bind     2733..2752
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2885..3109
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3098..3115
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(3156..3701)
                     /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             complement(4458..4487)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             complement(4497..4526)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      4526..4535
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(4629..4653)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    4663..4681
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4684..4702
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        complement(4704..4907)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4733..4753)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4908..5287
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5459..5478
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5553..5657
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5658..5726,5727..6455,1..63)
                     /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(5876..5895)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
       61 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
      121 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
      181 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
      241 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
      301 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
      361 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
      421 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
      481 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
      541 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
      601 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
      661 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
      721 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
      781 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
      841 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
      901 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
      961 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     1021 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     1081 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     1141 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     1201 tttcacacag gaaacagcta tgaccatgat tacgccaagc tctagctaga ggtcgacggt
     1261 atacagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     1321 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     1381 gcaataaaca agttggggtg ggcgaagaac tccagcatga gatccccgcg ctggaggatc
     1441 atccagccgg cgtcccggaa aacgattccg aagcccaacc tttcatagaa ggcggcggtg
     1501 gaatcgaaat ctcgtagtac gtgctattcc tttgccctcg gacgagtgct ggggcgtcgg
     1561 tttccactat cggcgagtac ttctacacag ccatcggtcc agacggccgc gcttctgcgg
     1621 gcgatttgtg tacgcccgac agtcccggct ccggatcgga cgattgcgtc gcatcgaccc
     1681 tgcgcccaag ctgcatcatc gaaattgccg tcaaccaagc tctgatagag ttggtcaaga
     1741 ccaatgcgga gcatatacgc ccggagccgc ggcgatcctg caagctccgg atgcctccgc
     1801 tcgaagtagc gcgtctgctg ctccatacaa gccaaccacg gcctccagaa gaagatgttg
     1861 gcgacctcgt attgggaatc cccgaacatc gcctcgctcc agtcaatgac cgctgttatg
     1921 cggccattgt ccgtcaggac attgttggag ccgaaatccg cgtgcacgag gtgccggact
     1981 tcggggcagt cctcggccca aagcatcagc tcatcgagag cctgcgcgac ggacgcactg
     2041 acggtgtcgt ccatcacagt ttgccagtga tacacatggg gatcagcaat cgcgcatatg
     2101 aaatcacgcc atgtagtgta ttgaccgatt ccttgcggtc cgaatgggcc gaacccgctc
     2161 gtctggctaa gatcggccgc agcgatcgca tccatggcct ccgcgaccgg ctgcagaaca
     2221 gcgggcagtt cggtttcagg caggtcttgc aacgtgacac cctgtgcacg gcgggagatg
     2281 caataggtca ggctctcgct gaattcccca atgtcaagca cttccggaat cgggagcgcg
     2341 gccgatgcaa agtgccgata aacataacga tctttgtaga aaccatcggc gcagctattt
     2401 acccgcagga catatccacg ccctcctaca tcgaagctga aagcacgaga ttcttcgccc
     2461 tccgagagct gcatcaggtc ggagacgctg tcgaactttt cgatcagaaa cttctcgaca
     2521 gacgtcgcgg tgagttcagg ctttttggcc aaggaagttc ctatactttc tagagaatag
     2581 gaacttcgga ataggaactt ctaggtacgt gaaccatcac cctaatcaag ttttttgggg
     2641 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga
     2701 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct
     2761 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat
     2821 gcgccgctac agggcgcgtg gggatacccc ctagagcccc agctggttct ttccgcctca
     2881 gaagccatag agcccaccgc atccccagca tgcctgctat tgtcttccca atcctccccc
     2941 ttgctgtcct gccccacccc accccccaga atagaatgac acctactcag acaatgcgat
     3001 gcaatttcct cattttatta ggaaaggaca gtgggagtgg caccttccag ggtcaaggaa
     3061 ggcacggggg aggggcaaac aacagatggc tggcaactag aaggcacagt cgaggctgat
     3121 cagcgggttt aaacgggccc tctagactcg agtcaaccca gcccaggctt gcccagtctg
     3181 tggccctcgt gggccagcag ccactctttc acggcgagcc cgccctcgta gccccccacg
     3241 tccaggtcgc cctgcaccac ccggtggcag gggatcagaa tgggcacggg atttccgctc
     3301 agggcggttt tcacggcggc ggtggcggcg ggattgccgg ccagggcggc caggtggctg
     3361 tagctgatga cctctccgaa cttcaccact ttcagcagtt tccacagcac ctggcgggta
     3421 aagctctcct gctggaacac tgggtggtgc agggctggca cagggaactc ctcgatggcc
     3481 tcaggctggt gaaagtaggc gttgagccag gcggtggcct gcatcagtgg ctctggtccg
     3541 cccagcacgg cggctggggc aggcacttcc acggcgtcgg cggcagatgt tcctttgccc
     3601 aggaagatga tacggtgcag gccctgttcg cacccagaca gttccagctt gcccagaggg
     3661 ctatccaggg tggtgcgctt catttcgcag tctttgtcca tgccaccaga tccggatccg
     3721 gaacctccac tgccataaat gcctgaattg gcggcgatag cgctaggtat cgggttacca
     3781 tccttgagca ggccttgcat agctttgaca atcaattcac aatctgaatt ggtagcgaag
     3841 atcggaatag taagctccat gttaaggtat gatcgccagg ctcccttggg gacttcaacc
     3901 tttatggtgt actttcggtt ttgtgcgctt gactggcgaa cactacacgt aactttataa
     3961 gcctgggagc gggaattaga actaatccat tctgcaatgc cgttagcaaa gttggaaggt
     4021 gccactgtca cgtcgcccgt gccaccattg tctacaagca caaactgggt gaagtttgaa
     4081 gccatggcat agatgccgga gtttgcggcg atggcgctag ggatagggtt tccatctttc
     4141 agcagacctt gcattgcctt aacgataagt tcacagtcgg agttggtggc gaagatggga
     4201 atagtgagtt ccatgttcag ataagatctc cacgcaccct ttggaacttc tactttgatg
     4261 gtatactttc gattttgggc ggagctttgt ctgacggaac acgttacttt atacgcttgt
     4321 gacctggaat tactggaaat ccactcggct attccgttgg caaaattgga gggagcgacc
     4381 gtaacgtccc ccgtaccgcc gttatccacc aagacgaact gagtaaaatt ggaggctcca
     4441 gaaccgcttc ctgaacccaa gtcttcctct gaaatcaatt tctgctctcc tgacccaaga
     4501 tcttcctcgc tgatgagttt ttgctccatg gtggcggcaa gcttaagttt aaacgctaga
     4561 gtccggaggc tggatcggtc ccggtgtctt ctatggaggt caaaacagcg tggatggcgt
     4621 ctccaggcga tctgacggtt cactaaacga gctcgtcgac gatctctatc actgataggg
     4681 agatctctat cactgatagg gagagctctg cttatataga cctcccaccg tacacgccta
     4741 ccgcccattt gcgtcaatgg ggcggagttg ttacgacatt ttggaaagtc ccgttgattt
     4801 tggtgccaaa acaaactccc attgacgtca atggggtgga gacttggaaa tccccgtgag
     4861 tcaaaccgct atccacgccc attgatgtac tgccaaaacc gcatcaccat ggtaatagcg
     4921 atgactaata cgtagatgta ctgccaagta ggaaagtccc ataaggtcat gtactgggca
     4981 taatgccagg cgggccattt accgtcattg acgtcaatag ggggcgtact tggcatatga
     5041 tacacttgat gtactgccaa gtgggcagtt taccgtaaat actccaccca ttgacgtcaa
     5101 tggaaagtcc ctattggcgt tactatggga acatacgtca ttattgacgt caatgggcgg
     5161 gggtcgttgg gcggtcagcc aggcgggcca tttaccgtaa gttatgtaac gcggaactcc
     5221 atatatgggc tatgaactaa tgaccccgta attgattact attaataact agtcaataat
     5281 caatgtcaac gcgtatatct ggcccgtaca tcgcgaagca gcgcaaaacg cctaacccta
     5341 agcagattct tcatgcaatt gtcggtcaag ccttgccttg ttgtagctta aattttgctc
     5401 gcgcactact cagcgacctc caacacacaa gcagggagca gatactggct taactatgcg
     5461 gcatcagagc agattgtact gagagtgcac cataggggat cgggagatct cccgatccgt
     5521 cgacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     5581 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     5641 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     5701 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     5761 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     5821 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     5881 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     5941 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     6001 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     6061 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     6121 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     6181 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     6241 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     6301 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     6361 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     6421 gtatcgtagt tatctacacg acggggagtc aggca
//
