LOCUS       sequence-389378-        8102 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CMV overexpression of ChemoL-NAD in mammalian cell lines.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8102)
  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 8102)
  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-ChemoL-NAD
FEATURES             Location/Qualifiers
     source          1..8102
                     /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(1470..1472,1473..1985)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIE"
     primer_bind     complement(1512..1533)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1773..1792)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2120..2141
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4054..4071)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4060..4284
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4417..4436)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4568..4615
                     /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             4623..5645
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4623..5645
                     /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    5775..5896
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5812..5831)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5866..5885
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5945..5961)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5945..5961)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5958..5980)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5969..5985
                     /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(5993..5999,6000..6017,6018..6023))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6038..6059
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6176..6193)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6347..6935)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6427..6446)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7106..7897,7898..7966))
                     /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     7729..7748
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7967..8071)
                     /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 aagaagctgc
      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 gtctgaggcg gcgaccaaat gggccagatc gaaaaaattt
      961 ttaaggtggt gtaccctgtg gatgatcatc actttaaggt gatcctgcac tatggcacac
     1021 tggtaatcga cggggttacg ccgaacatga tcgactattt cggacggccg tatgaaggca
     1081 tcgccgtgtt cgacggcaaa aagatcactg taacagggac cctgtggaac ggcaacaaaa
     1141 ttatcgacga gcgcctgatc aaccccgacg gctccctgct gttccgagta accatcaacg
     1201 gagtgaccgg ctggcggctg tgcgaacgca ttctggcggg tggcaccggt ggctctggcg
     1261 gtaccggtgg ctctatggtc ttcacactcg aagatttcgt tggggactgg cgacagacag
     1321 ccggctacaa cctggaccaa gtccttgaac agggaggtgt gtccagtttg tttcagaatc
     1381 tcggggtgtc cgtaactccg atccaaagga ttgtcctgag cggtgaaaat gggctgaaga
     1441 tcgacatcca tgtcatcatc ccgtatgaag tgagcaaggg cgaggagctg ttcaccgggg
     1501 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtatccg
     1561 gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg
     1621 gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct
     1681 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag
     1741 gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg
     1801 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca
     1861 aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct
     1921 atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca
     1981 tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg
     2041 gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccaaactg agcaaagacc
     2101 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gcgcgccgcc gggatcactg
     2161 gtggcaccat gacactggaa gaagctcgta aacgtgtaaa cgaacttcgt gatctgatcc
     2221 gttatcacaa ttaccggtac tacgtgttgg ccgaccccga gattagcgat gcagagtacg
     2281 atcgtctgct gcgcgaactg aaggagttgg aggaacgttt tcctgagctc aaatcgccgg
     2341 attcaccgac attacaagtg ggcgcacggc cgctggaagc cacgttccgt cccgtgcgcc
     2401 atccgacacg catgtacagt ctggataacg cattcaactt ggatgaactg aaagcctttg
     2461 aggaacgtat cgaacgcgcg ctcgggcgca aaggtccgtt cgcctataca gttgaacatc
     2521 tagtggatgg cctgagcgtc aacttatatt acgaagaagg agtgttggtt tacggcgcaa
     2581 cgcgtggcga cggtgaagtg ggtgaggaag ttactcaaaa tttactcacc atcccgacta
     2641 ttccacgccg tctgaaaggt gtcccggaac gcctcgaggt gcgtggcgaa gtgtacatgc
     2701 cgattgaggc gtttctgcgc ctgaatgaag aactggagga acgtggcgag cgcattttta
     2761 aaaatccgcg taacgctgca gcaggcagct tgcgccagaa agatccgcgc attacggcta
     2821 agcgtggttt gcgcgcaacc ttctgggcgt taggtctggg tctcgaagag gtggagcgtg
     2881 aaggagtagc gactcaattt gcgttgctgc attggctcaa agagaaaggg ttcccggtcg
     2941 aacacggata tgcacgtgcc gttggggcgg aaggcgtgga ggcggtttat caggactggc
     3001 tgaaaaaacg ccgtgcgctg ccgtttgagg cgaatggagt tgcggtcaaa ctggatgaac
     3061 ttgccctgtg gcgtgaactg ggatacaccg cgcgtgcacc gcgttttgcc attgcgtaca
     3121 agttcccttc tggcggtatc ggtactggct ttccattcga cccccattat gtggaagtcc
     3181 tgggcgagcg catgcactac gtcgatgttg gtccgcgcga tggcacccct gtgctgttcc
     3241 tgcacggtaa cccgacctcc tcctacgtgt ggcgcaacat catcccgcat gttgcaccga
     3301 cccatcgctg cattgctcca gacctgatcg gtatgggcaa atccgacaaa ccagacctgg
     3361 gttatttctt cgacgaccac gtccgcttca tggatgcctt catcgaagcc ctgggtctgg
     3421 aagaggtcgt cctggtcatt cacgactggg gctccgctct gggtttccac tgggccaagc
     3481 gcaatccaga gcgcgtcaaa ggtattgcat ttatggagtt catccgccct atcccgacct
     3541 gggacgaatg gccagaattt gcccgcgaga ccttccaggc cttccgcacc accgacgtcg
     3601 gccgcaagct gatcatcgat cagaacgttt ttatcgaggg tacgctgccg atgggtgtcg
     3661 tccgcccgct gactgaagtc gagatggacc attaccgcga gccgttcctg aatcctgttg
     3721 accgcgagcc actgtggcgc ttcccaaacg agctgccaat cgccggtgag ccagcgaaca
     3781 tcgtcgcgct ggtcgaagaa tacatggact ggctgcacca gtcccctgtc ccgaagctgc
     3841 tgttctgggg caccccaggc gttctgatcc caccggccga agccgctcgc ctggccaaaa
     3901 gcctgcctaa ctgcaaggct gtggacatcg gcccgggtga aaatctgctg caagaagaca
     3961 acccggacct gatcggcagc gagatcgcgc gctggctgtc tactctggag atttccggtt
     4021 agtctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     4081 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     4141 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     4201 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     4261 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     4321 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     4381 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     4441 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     4501 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     4561 gtacctagaa gttcctattc cgaagttcct attctctaga aagtatagga acttccttgg
     4621 ccaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca
     4681 gcgtctccga cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg
     4741 taggagggcg tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc
     4801 gttatgttta tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg
     4861 gggaattcag cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc
     4921 aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg
     4981 cgatcgctgc ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa
     5041 tcggtcaata cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc
     5101 actggcaaac tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc
     5161 tgatgctttg ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct
     5221 ccaacaatgt cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga
     5281 tgttcgggga ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt
     5341 gtatggagca gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc
     5401 ggctccgggc gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg
     5461 gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag
     5521 ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct
     5581 gtgtagaagt actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg
     5641 aatagcacgt actacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     5701 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     5761 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     5821 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     5881 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
     5941 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     6001 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     6061 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     6121 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     6181 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     6241 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     6301 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     6361 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     6421 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     6481 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     6541 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     6601 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     6661 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     6721 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     6781 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     6841 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     6901 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     6961 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     7021 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     7081 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     7141 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     7201 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     7261 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     7321 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     7381 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     7441 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     7501 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     7561 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     7621 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     7681 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     7741 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     7801 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     7861 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     7921 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     7981 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     8041 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     8101 tc
//
