LOCUS       sequence-393562-        7552 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV overexpression of ChemoG-NAD in mammalian cell lines.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7552)
  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 7552)
  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-NAD
FEATURES             Location/Qualifiers
     source          1..7552
                     /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     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
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     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"
     primer_bind     complement(3504..3521)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3510..3734
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3867..3886)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4018..4065
                     /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             4073..5095
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4073..5095
                     /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    5225..5346
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5262..5281)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5316..5335
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5395..5411)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5395..5411)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5408..5430)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5419..5435
                     /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(5443..5449,5450..5467,5468..5473))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5488..5509
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5626..5643)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5797..6385)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5877..5896)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6556..7347,7348..7416))
                     /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     7179..7198
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7417..7521)
                     /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 acaactccga 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 agcgtatccg 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 cgagggcgac 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 gcgcgccgcc gggatcactg gtggcaccat
     1621 gacactggaa gaagctcgta aacgtgtaaa cgaacttcgt gatctgatcc gttatcacaa
     1681 ttaccggtac tacgtgttgg ccgaccccga gattagcgat gcagagtacg atcgtctgct
     1741 gcgcgaactg aaggagttgg aggaacgttt tcctgagctc aaatcgccgg attcaccgac
     1801 attacaagtg ggcgcacggc cgctggaagc cacgttccgt cccgtgcgcc atccgacacg
     1861 catgtacagt ctggataacg cattcaactt ggatgaactg aaagcctttg aggaacgtat
     1921 cgaacgcgcg ctcgggcgca aaggtccgtt cgcctataca gttgaacatc tagtggatgg
     1981 cctgagcgtc aacttatatt acgaagaagg agtgttggtt tacggcgcaa cgcgtggcga
     2041 cggtgaagtg ggtgaggaag ttactcaaaa tttactcacc atcccgacta ttccacgccg
     2101 tctgaaaggt gtcccggaac gcctcgaggt gcgtggcgaa gtgtacatgc cgattgaggc
     2161 gtttctgcgc ctgaatgaag aactggagga acgtggcgag cgcattttta aaaatccgcg
     2221 taacgctgca gcaggcagct tgcgccagaa agatccgcgc attacggcta agcgtggttt
     2281 gcgcgcaacc ttctgggcgt taggtctggg tctcgaagag gtggagcgtg aaggagtagc
     2341 gactcaattt gcgttgctgc attggctcaa agagaaaggg ttcccggtcg aacacggata
     2401 tgcacgtgcc gttggggcgg aaggcgtgga ggcggtttat caggactggc tgaaaaaacg
     2461 ccgtgcgctg ccgtttgagg cgaatggagt tgcggtcaaa ctggatgaac ttgccctgtg
     2521 gcgtgaactg ggatacaccg cgcgtgcacc gcgttttgcc attgcgtaca agttcccttc
     2581 tggcggtatc ggtactggct ttccattcga cccccattat gtggaagtcc tgggcgagcg
     2641 catgcactac gtcgatgttg gtccgcgcga tggcacccct gtgctgttcc tgcacggtaa
     2701 cccgacctcc tcctacgtgt ggcgcaacat catcccgcat gttgcaccga cccatcgctg
     2761 cattgctcca gacctgatcg gtatgggcaa atccgacaaa ccagacctgg gttatttctt
     2821 cgacgaccac gtccgcttca tggatgcctt catcgaagcc ctgggtctgg aagaggtcgt
     2881 cctggtcatt cacgactggg gctccgctct gggtttccac tgggccaagc gcaatccaga
     2941 gcgcgtcaaa ggtattgcat ttatggagtt catccgccct atcccgacct gggacgaatg
     3001 gccagaattt gcccgcgaga ccttccaggc cttccgcacc accgacgtcg gccgcaagct
     3061 gatcatcgat cagaacgttt ttatcgaggg tacgctgccg atgggtgtcg tccgcccgct
     3121 gactgaagtc gagatggacc attaccgcga gccgttcctg aatcctgttg accgcgagcc
     3181 actgtggcgc ttcccaaacg agctgccaat cgccggtgag ccagcgaaca tcgtcgcgct
     3241 ggtcgaagaa tacatggact ggctgcacca gtcccctgtc ccgaagctgc tgttctgggg
     3301 caccccaggc gttctgatcc caccggccga agccgctcgc ctggccaaaa gcctgcctaa
     3361 ctgcaaggct gtggacatcg gcccgggtga aaatctgctg caagaagaca acccggacct
     3421 gatcggcagc gagatcgcgc gctggctgtc tactctggag atttccggtt agtctagagg
     3481 gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     3541 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     3601 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     3661 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc
     3721 ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca
     3781 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     3841 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     3901 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     3961 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtacctagaa
     4021 gttcctattc cgaagttcct attctctaga aagtatagga acttccttgg ccaaaaagcc
     4081 tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga
     4141 cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg
     4201 tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta
     4261 tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattcag
     4321 cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc
     4381 tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc
     4441 ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata
     4501 cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac
     4561 tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg
     4621 ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt
     4681 cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga
     4741 ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca
     4801 gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc
     4861 gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga
     4921 tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt
     4981 cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct gtgtagaagt
     5041 actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg aatagcacgt
     5101 actacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     5161 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     5221 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     5281 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5341 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     5401 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5461 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5521 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5581 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5641 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5701 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5761 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5821 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5881 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5941 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     6001 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     6061 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     6121 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     6181 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     6241 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6301 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6361 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6421 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6481 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6541 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6601 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6661 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6721 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6781 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6841 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6901 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6961 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     7021 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     7081 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     7141 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7201 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7261 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7321 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7381 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7441 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7501 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
