LOCUS       sequence-381763-        7222 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV overexpression of ChemoG-CaM in mammalian cell lines.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7222)
  AUTHORS   Hellweg L, Edenhofer A, Barck L, Huppertz MC, Frei MS, Tarnawski M, 
            Bergner A, Koch B, Johnsson K, Hiblot J
  TITLE     A general method for the development of multicolor biosensors with 
            large dynamic ranges.
  JOURNAL   Nat Chem Biol. 2023 Jun 8. doi: 10.1038/s41589-023-01350-1.
  PUBMED    37291200
REFERENCE   2  (bases 1 to 7222)
  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: "Nat Chem 
            Biol. 2023 Jun 8. doi: 10.1038/s41589-023-01350-1."
COMMENT     Sequence Label: pCDNA5/FRT-ChemoG-CaM
FEATURES             Location/Qualifiers
     source          1..7222
                     /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"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(917..919,920..922,923..1435)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFDGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIE"
     primer_bind     complement(962..983)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1223..1242)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1570..1591
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1625..2065
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
                     ELQDMINEVDADGDGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYISAAEL
                     RHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVVMMTAK"
     primer_bind     complement(3174..3191)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3180..3404
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3537..3556)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3688..3735
                     /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             3743..4765
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3743..4765
                     /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    4895..5016
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4932..4951)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4986..5005
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5065..5081)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5065..5081)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5078..5100)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5089..5105
                     /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(5113..5119,5120..5137,5138..5143))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5158..5179
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5296..5313)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5467..6055)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5547..5566)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6226..7017,7018..7086))
                     /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     6849..6868
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7087..7191)
                     /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 ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt accaccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
      961 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     1021 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     1081 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     1141 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     1201 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     1261 cgatggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     1321 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
     1381 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
     1441 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     1501 gctgcccgac aaccactacc tgagcaccca gtccaaactg agcaaagacc ccaacgagaa
     1561 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactg gtggcaccct
     1621 gccggaccaa ctgactgaag agcagatcgc agaatttaaa gaggctttct ccctatttga
     1681 caaggacggg gatgggacaa taacaaccaa ggagctgggg acggtgatgc ggtctctggg
     1741 gcagaacccc acagaagcag agctgcagga catgatcaat gaagtagatg ccgacggtga
     1801 cggcacaatc gacttccctg agttcctgac aatgatggca agaaaaatga aagacacaga
     1861 cagtgaagaa gaaattagag aagcgttccg tgtgtttgat aaggatggca atggctacat
     1921 cagtgcagca gagcttcgcc acgtgatgac aaaccttgga gagaagttaa cagatgaaga
     1981 ggttgatgaa atgatcaggg aagcagacat cgatggggat ggtcaggtaa actacgaaga
     2041 gtttgtagtg atgatgacag cgaaggaatt ccctcctcct ccgccaccgc caccaccgcc
     2101 gccgccgcca ccaccacctc ctccgcctcc tccaccacca ccgccaccgc ctccaccacc
     2161 tggcggcagc agggtcgact catcacgtcg taagttcaat aagacaggta aggcactgag
     2221 agctataggt cggctgagct cactcgagtc tggcggtatc ggtactggct ttccattcga
     2281 cccccattat gtggaagtcc tgggcgagcg catgcactac gtcgatgttg gtccgcgcga
     2341 tggcacccct gtgctgttcc tgcacggtaa cccgacctcc tcctacgtgt ggcgcaacat
     2401 catcccgcat gttgcaccga cccatcgctg cattgctcca gacctgatcg gtatgggcaa
     2461 atccgacaaa ccagacctgg gttatttctt cgacgaccac gtccgcttca tggatgcctt
     2521 catcgaagcc ctgggtctgg aagaggtcgt cctggtcatt cacgactggg gctccgctct
     2581 gggtttccac tgggccaagc gcaatccaga gcgcgtcaaa ggtattgcat ttatggagtt
     2641 catccgccct atcccgacct gggacgaatg gccagaattt gcccgcgaga ccttccaggc
     2701 cttccgcacc accgacgtcg gccgcaagct gatcatcgat cagaacgttt ttatcgaggg
     2761 tacgctgccg atgggtgtcg tccgcccgct gactgaagtc gagatggacc attaccgcga
     2821 gccgttcctg aatcctgttg accgcgagcc actgtggcgc ttcccaaacg agctgccaat
     2881 cgccggtgag ccagcgaaca tcgtcgcgct ggtcgaagaa tacatggact ggctgcacca
     2941 gtcccctgtc ccgaagctgc tgttctgggg caccccaggc gttctgatcc caccggccga
     3001 agccgctcgc ctggccaaaa gcctgcctaa ctgcaaggct gtggacatcg gcccgggtga
     3061 aaatctgctg caagaagaca acccggacct gatcggcagc gagatcgcgc gctggctgtc
     3121 tactctggag atttccggtt agtctagagg gcccgtttaa acccgctgat cagcctcgac
     3181 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     3241 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     3301 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     3361 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     3421 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     3481 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     3541 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     3601 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     3661 tgattagggt gatggttcac gtacctagaa gttcctattc cgaagttcct attctctaga
     3721 aagtatagga acttccttgg ccaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt
     3781 tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg gcgaagaatc
     3841 tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa atagctgcgc
     3901 cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg cgctcccgat
     3961 tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca tctcccgccg
     4021 tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc
     4081 ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga gcgggttcgg
     4141 cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca tatgcgcgat
     4201 tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca gtgcgtccgt
     4261 cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag tccggcacct
     4321 cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca taacagcggt
     4381 cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca acatcttctt
     4441 ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc ggaggcatcc
     4501 ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc ttgaccaact
     4561 ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga
     4621 cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc
     4681 ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc gacgccccag
     4741 cactcgtccg agggcaaagg aatagcacgt actacgagat ttcgattcca ccgccgcctt
     4801 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     4861 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     4921 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     4981 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     5041 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     5101 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     5161 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     5221 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     5281 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     5341 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     5401 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     5461 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5521 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5581 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5641 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5701 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5761 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5821 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5881 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     5941 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     6001 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     6061 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     6121 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     6181 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     6241 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     6301 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     6361 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     6421 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6481 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6541 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6601 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     6661 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     6721 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     6781 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     6841 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     6901 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     6961 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     7021 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     7081 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     7141 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     7201 ccgaaaagtg ccacctgacg tc
//
