LOCUS       sequence-355921-        5980 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV overexpression of SNAP-tag fast at the outer leaflet of the 
            plasma membrane of mammalian cell lines.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5980)
  AUTHORS   Frei MS, Koch B, Hiblot J, Johnsson K
  TITLE     Live-Cell Fluorescence Lifetime Multiplexing Using Synthetic 
            Fluorescent Probes.
  JOURNAL   ACS Chem Biol. 2022 Jun 17;17(6):1321-1327. doi: 
            10.1021/acschembio.2c00041. Epub 2022 May 18.
  PUBMED    35584304
REFERENCE   2  (bases 1 to 5980)
  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: 
            "10.1021/acschembio.2c00041"; journalName: "ACS Chem Biol"; date: 
            "2022-06-17- 17"; volume: "17"; issue: "6"; pages: "1321-1327"
COMMENT     Sequence Label: pcDNA5/FRT/TO_IgKchL_HA_STf_myc_PDGFRtmb
FEATURES             Location/Qualifiers
     source          1..5980
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     sig_peptide     1043..1102
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             1106..1132
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1706..1735
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1739..1885
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(1932..1949)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1938..2162
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2295..2314)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2446..2493
                     /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             2501..3523
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2501..3523
                     /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    3653..3774
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3690..3709)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3744..3763
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3823..3839)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3823..3839)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3836..3858)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3847..3863
                     /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(3871..3877,3878..3895,3896..3901))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3916..3937
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4054..4071)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4225..4813)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4305..4324)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4984..5775,5776..5844))
                     /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     5607..5626
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5845..5949)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cactagtcca gtgtggtgga
     1021 attctgcaga tatccagcaa ccatggagac agacacactc ctgctatggg tactgctgct
     1081 ctgggttcca ggttccactg gtgactatcc atatgatgtt ccagattatg ctggggccca
     1141 gccggccaga tcagataaag actgcgaaat gaaacgcact accctggaca gcccgctggg
     1201 taaactggaa ctgagcggtt gcgagcaggg cctgcaccgc atcattttcc tgggcaaagg
     1261 tacgagcgct gcggatgcgg tcgaagtacc ggctccggct gcggttctgg gtggcccgga
     1321 gccgctgatg caggctaccg catggctgaa cgcatatttt caccagccag aggcgattga
     1381 agagttccct gtaccggccc tgcatcaccc ggtatttcag caggaatcct ttacccgtca
     1441 ggtactgtgg aaactgctga aagtagtgaa attcggcgag gtgatctcct attcccatct
     1501 ggcggcgctg gctggcaacc cagcggcaac tgcggcggtg aaaaccgctc tgtctggtaa
     1561 tccggtgcca attctgatcc cgtgccaccg tgtggttcag ggcgacctgg acgtaggtgg
     1621 ctacgaaggc ggcctggctg tgaaagaatg gctgctggcg cacgaaggtc accgtctggg
     1681 taaaccaggc ctgggcggcg tcgacgaaca aaaactcatc tcagaagagg atctgaatgc
     1741 tgtgggccag gacacgcagg aggtcatcgt ggtgccacac tccttgccct ttaaggtggt
     1801 ggtgatctca gccatcctgg ccctggtggt gctcaccatc atctccctta tcatcctcat
     1861 catgctttgg cagaagaagc cacgttagta accgctcgag tctagagggc ccgtttaaac
     1921 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     1981 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     2041 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     2101 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     2161 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     2221 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     2281 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     2341 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     2401 cctcgacccc aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg
     2461 aagttcctat tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc
     2521 gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct
     2581 ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct
     2641 gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc
     2701 atcggccgcg ctcccgattc cggaagtgct tgacattggg gaattcagcg agagcctgac
     2761 ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact
     2821 gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg ccgatcttag
     2881 ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg
     2941 tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga
     3001 caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg
     3061 ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa
     3121 tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga
     3181 ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta
     3241 cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt atatgctccg
     3301 cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg
     3361 ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca
     3421 aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt gtagaagtac tcgccgatag
     3481 tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt
     3541 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     3601 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     3661 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     3721 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     3781 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     3841 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     3901 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     3961 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4021 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4081 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4141 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4201 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4261 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4321 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4381 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4441 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4501 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4561 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4621 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     4681 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4741 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4801 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     4861 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     4921 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     4981 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5041 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5101 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5161 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5221 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5281 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5341 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5401 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5461 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5521 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5581 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5641 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     5701 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5761 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     5821 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     5881 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     5941 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
//
