LOCUS       sequence-300659-        7222 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a G alpha subunit (GNAO1 Isoform Alpha 1) containing RLuc8 
            as an optimal component of a BRET2 biosensor for studying 
            heterotrimeric G protein dissociation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7222)
  AUTHORS   Olsen RHJ, DiBerto JF, English JG, Glaudin AM, Krumm BE, Slocum ST, 
            Che T, Gavin AC, McCorvy JD, Roth BL, Strachan RT
  TITLE     TRUPATH, an open-source biosensor platform for interrogating the 
            GPCR transducerome.
  JOURNAL   Nat Chem Biol. 2020 May 4. pii: 10.1038/s41589-020-0535-8. doi: 
            10.1038/s41589-020-0535-8.
  PUBMED    32367019
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. 2020 May 4. pii: 10.1038/s41589-020-0535-8. doi: 
            10.1038/s41589-020-0535-8."
COMMENT     Sequence Label: pcDNA5/FRT/TO-GAlphaoA-RLuc8
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"
     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"
     CDS             1392..2324
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNATSSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGAALAFHYAYEHQDRIKAIVHMESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETVLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKLFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFLQEDAPDEMGKYIKSFVERVLKNEQ"
     primer_bind     complement(3175..3192)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3181..3405
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3538..3557)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3689..3736
                     /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)."
     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 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 tatcaccacc gccagctcct ccctggccaa ggacacttca tcgaccggtg
     1081 agaacctgta cttccagcta atgggatgta ctctgagcgc agaggagaga gccgccctcg
     1141 agcggagcaa ggcgattgag aaaaacctca aagaggatgg catcagcgcc gccaaagacg
     1201 tgaaattact cctgctcggg gctggagaat caggaaaaag caccattgtg aagcagatga
     1261 agatcatcca tgaagatggc ttctccggag aagacgtgaa acagtacaag cctgttgtct
     1321 acagcaacac tatccagtcc ctggcagcca tcgtccgggc catggacact ttgtctggtg
     1381 gtggtggatc catggcttcc aaggtgtacg accccgagca acgcaaacgc atgatcactg
     1441 ggcctcagtg gtgggctcgc tgcaagcaaa tgaacgtgct ggactccttc atcaactact
     1501 atgattccga gaagcacgcc gagaacgccg tgatttttct gcatggtaac gctacctcca
     1561 gctacctgtg gaggcacgtc gtgcctcaca tcgagcccgt ggctagatgc atcatccctg
     1621 atctgatcgg aatgggtaag tccggcaaga gcgggaatgg ctcatatcgc ctcctggatc
     1681 actacaagta cctcaccgct tggttcgagc tgctgaacct tccaaagaaa atcatctttg
     1741 tgggccacga ctggggggct gctctggcct ttcactacgc ctacgagcac caagacagga
     1801 tcaaggccat cgtccatatg gagagtgtcg tggacgtgat cgagtcctgg gacgagtggc
     1861 ctgacatcga ggaggatatc gccctgatca agagcgaaga gggcgagaaa atggtgcttg
     1921 agaataactt cttcgtcgag accgtgctcc caagcaagat catgcggaaa ctggagcctg
     1981 aggagttcgc tgcctacctg gagccattca aggagaaggg cgaggttaga cggcctaccc
     2041 tctcctggcc tcgcgagatc cctctcgtta agggaggcaa gcccgacgtc gtccagattg
     2101 tccgcaacta caacgcctac cttcgggcca gcgacgatct gcctaagctg ttcatcgagt
     2161 ccgaccctgg gttcttttcc aacgctattg tcgagggagc taagaagttc cctaacaccg
     2221 agttcgtgaa ggtgaagggc ctccacttcc tccaggagga cgctccagat gaaatgggta
     2281 agtacatcaa gagcttcgtg gagcgcgtgc tgaagaacga gcagagcgga ggaggcggca
     2341 gtggcatcga atatggtgat aaggagagaa aggctgacgc caagatggtg tgtgatgtgg
     2401 tgagtcggat ggaagacacc gagcccttct ctgcagagct gctttctgcc atgatgcggc
     2461 tctggggcga ctcaggaatc caagagtgct tcaaccggtc ccgggagtat cagctcaacg
     2521 actctgccaa atactacctg gacagcctgg atcggattgg ggccgccgac taccagccca
     2581 ccgagcagga catcctccga accagggtca aaaccactgg catcgtagaa acccacttca
     2641 cattcaagaa cctccacttc aggctgtttg acgtcggagg ccagcgatct gaacgcaaga
     2701 agtggatcca ttgcttcgag gacgtcacgg ccatcatttt ctgtgtcgcg ctcagcggct
     2761 atgaccaggt gctccacgaa gacgaaacca cgaaccgcat gcacgagtct ctcatgctct
     2821 tcgactccat ctgtaacaac aagttcttca tcgatacctc catcattctc ttcctcaaca
     2881 agaaagatct ctttggcgag aagatcaaga agtcaccttt gaccatctgc tttcctgaat
     2941 acacaggccc caatacctat gaagacgcag ccgcctacat ccaagcacaa tttgaaagca
     3001 aaaaccgctc acccaacaaa gaaatatatt gtcacatgac ttgtgccaca gacacgaata
     3061 acatccaggt ggtgttcgac gccgtcaccg acatcatcat tgccaacaac ctccggggct
     3121 gcggcttgta ctgatagctc gagtctagag ggcccgttta aacccgctga tcagcctcga
     3181 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     3241 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     3301 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     3361 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     3421 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     3481 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     3541 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     3601 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     3661 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag
     3721 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt
     3781 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat
     3841 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg
     3901 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga
     3961 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc
     4021 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc
     4081 catcgtccgg 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
//
