LOCUS       sequence-356088-        6142 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV overexpression of SNAP-tag fast at the mitochondria surface of 
            mammalian cell lines.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6142)
  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 6142)
  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_TOMM20-STf
FEATURES             Location/Qualifiers
     source          1..6142
                     /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..613
                     /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"
     primer_bind     complement(2094..2111)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2100..2324
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2457..2476)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2608..2655
                     /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             2663..3685
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2663..3685
                     /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    3815..3936
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3852..3871)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3906..3925
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3985..4001)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3985..4001)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3998..4020)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4009..4025
                     /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(4033..4039,4040..4057,4058..4063))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4078..4099
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4216..4233)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4387..4975)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4467..4486)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5146..5937,5938..6006))
                     /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     5769..5788
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6007..6111)
                     /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 cattgtgatg 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 ccatggtggg tcggaacagc gccatcgccg ccggtgtatg
     1081 cggggccctt ttcattgggt actgcatcta cttcgaccgc aaaagacgaa gtgaccccaa
     1141 cttcaagaac aggcttcgag aacgaagaaa gaaacagaag cttgccaagg agagagctgg
     1201 gctttccaag ttacctgacc ttaaagatgc tgaagctgtt cagaagttct tccttgaaga
     1261 aatacagctt ggtgaagagt tactagctca aggtgaatat gagaagggcg tagaccatct
     1321 gacaaatgca attgctgtgt gtggacagcc acagcagtta ctgcaggtct tacagcaaac
     1381 tcttccacca ccagtgttcc agatgcttct gactaagctc ccaacaatta gtcagagaat
     1441 tgtaagtgct cagagcttgg ctgaagatga tgtggaaggc ggtagcgggg atccaccggt
     1501 cggaggtgat aaagactgcg aaatgaaacg cactaccctg gacagcccgc tgggtaaact
     1561 ggaactgagc ggttgcgagc agggcctgca ccgcatcatt ttcctgggca aaggtacgag
     1621 cgctgcggat gcggtcgaag taccggctcc ggctgcggtt ctgggtggcc cggagccgct
     1681 gatgcaggct accgcatggc tgaacgcata ttttcaccag ccagaggcga ttgaagagtt
     1741 ccctgtaccg gccctgcatc acccggtatt tcagcaggaa tcctttaccc gtcaggtact
     1801 gtggaaactg ctgaaagtag tgaaattcgg cgaggtgatc tcctattccc atctggcggc
     1861 gctggctggc aacccagcgg caactgcggc ggtgaaaacc gctctgtctg gtaatccggt
     1921 gccaattctg atcccgtgcc accgtgtggt tcagggcgac ctggacgtag gtggctacga
     1981 aggcggcctg gctgtgaaag aatggctgct ggcgcacgaa ggtcaccgtc tgggtaaacc
     2041 aggcctgggc taaccgctcg agtctagagg gcccgtttaa acccgctgat cagcctcgac
     2101 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     2161 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     2221 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     2281 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     2341 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     2401 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     2461 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     2521 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     2581 tgattagggt gatggttcac gtacctagaa gttcctattc cgaagttcct attctctaga
     2641 aagtatagga acttccttgg ccaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt
     2701 tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg gcgaagaatc
     2761 tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa atagctgcgc
     2821 cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg cgctcccgat
     2881 tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca tctcccgccg
     2941 tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc
     3001 ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga gcgggttcgg
     3061 cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca tatgcgcgat
     3121 tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca gtgcgtccgt
     3181 cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag tccggcacct
     3241 cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca taacagcggt
     3301 cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca acatcttctt
     3361 ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc ggaggcatcc
     3421 ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc ttgaccaact
     3481 ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga
     3541 cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc
     3601 ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc gacgccccag
     3661 cactcgtccg agggcaaagg aatagcacgt actacgagat ttcgattcca ccgccgcctt
     3721 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     3781 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     3841 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     3901 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     3961 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     4021 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     4081 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     4141 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     4201 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     4261 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     4321 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     4381 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     4441 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     4501 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     4561 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     4621 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     4681 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     4741 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     4801 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     4861 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     4921 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     4981 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     5041 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     5101 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     5161 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     5221 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     5281 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     5341 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     5401 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     5461 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     5521 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     5581 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     5641 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     5701 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     5761 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     5821 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     5881 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     5941 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     6001 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     6061 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     6121 ccgaaaagtg ccacctgacg tc
//
