LOCUS       sequence-300775-        7283 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a G alpha subunit (GNA15) 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 7283)
  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 7283)
  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-GAlpha15-RLuc8
FEATURES             Location/Qualifiers
     source          1..7283
                     /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             1851..2783
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNATSSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGAALAFHYAYEHQDRIKAIVHMESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETVLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKLFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFLQEDAPDEMGKYIKSFVERVLKNEQ"
     primer_bind     complement(3235..3252)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3241..3465
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3598..3617)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3749..3796
                     /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             3804..4826
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3804..4826
                     /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    4956..5077
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4993..5012)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5047..5066
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5126..5142)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5126..5142)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5139..5161)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5150..5166
                     /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(5174..5180,5181..5198,5199..5204))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5219..5240
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5357..5374)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5528..6116)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5608..5627)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6287..7078,7079..7147))
                     /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     6910..6929
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7148..7252)
                     /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 atggcccgct cgctgacctg gcgctgctgc ccctggtgcc
     1141 tgacggagga tgagaaggcc gccgcccggg tggaccagga gatcaacagg atcctcttgg
     1201 agcagaagaa gcaggaccgc ggggagctga agctgctgct tttgggccca ggcgagagcg
     1261 ggaagagcac cttcatcaag cagatgcgga tcatccacgg cgccggctac tcggaggagg
     1321 agcgcaaggg cttccggccc ctggtctacc agaacatctt cgtgtccatg cgggccatga
     1381 tcgaggccat ggagcggctg cagattccat tcagcaggcc cgagagcaag caccacgcta
     1441 gcctggtcat gagccaggac ccctataaag tgaccacgtt tgagaagcgc tacgctgcgg
     1501 ccatgcagtg gctgtggagg gatgccggca tccgggcctg ctatgagcgt cggcgggaat
     1561 tccacctgct cgattcagcc gtgtactacc tgtcccacct ggagcgcatc accgaggagg
     1621 gctacgtccc cacagctcag gacgtgctcc gcagccgcat gcccaccact ggcatcaacg
     1681 agtactgctt ctccgtgcag aaaaccaacc tgcggatcgt ggacgtcggg ggccagaagt
     1741 cagagcgtaa gaaatggatc cattgtttcg agaacgtgat cgccctcatc tacctggcct
     1801 cactgagtga atacgaccag tgcctggagg agtctggtgg tggtggatcc atggcttcca
     1861 aggtgtacga ccccgagcaa cgcaaacgca tgatcactgg gcctcagtgg tgggctcgct
     1921 gcaagcaaat gaacgtgctg gactccttca tcaactacta tgattccgag aagcacgccg
     1981 agaacgccgt gatttttctg catggtaacg ctacctccag ctacctgtgg aggcacgtcg
     2041 tgcctcacat cgagcccgtg gctagatgca tcatccctga tctgatcgga atgggtaagt
     2101 ccggcaagag cgggaatggc tcatatcgcc tcctggatca ctacaagtac ctcaccgctt
     2161 ggttcgagct gctgaacctt ccaaagaaaa tcatctttgt gggccacgac tggggggctg
     2221 ctctggcctt tcactacgcc tacgagcacc aagacaggat caaggccatc gtccatatgg
     2281 agagtgtcgt ggacgtgatc gagtcctggg acgagtggcc tgacatcgag gaggatatcg
     2341 ccctgatcaa gagcgaagag ggcgagaaaa tggtgcttga gaataacttc ttcgtcgaga
     2401 ccgtgctccc aagcaagatc atgcggaaac tggagcctga ggagttcgct gcctacctgg
     2461 agccattcaa ggagaagggc gaggttagac ggcctaccct ctcctggcct cgcgagatcc
     2521 ctctcgttaa gggaggcaag cccgacgtcg tccagattgt ccgcaactac aacgcctacc
     2581 ttcgggccag cgacgatctg cctaagctgt tcatcgagtc cgaccctggg ttcttttcca
     2641 acgctattgt cgagggagct aagaagttcc ctaacaccga gttcgtgaag gtgaagggcc
     2701 tccacttcct ccaggaggac gctccagatg aaatgggtaa gtacatcaag agcttcgtgg
     2761 agcgcgtgct gaagaacgag cagagcggag gaggcggcag taacaaccag gagaaccgca
     2821 tgaaggagag cctcgcattg tttgggacta tcctggaact accctggttc aaaagcacat
     2881 ccgtcatcct ctttctcaac aaaaccgaca tcctggagga gaaaatcccc acctcccacc
     2941 tggctaccta tttccccagt ttccagggcc ctaagcagga tgctgaggca gccaagaggt
     3001 tcatcctgga catgtacacg aggatgtaca ccgggtgcgt ggacggcccc gagggcagca
     3061 agaagggcgc acgatcccga cgcctcttca gccactacac atgtgccaca gacacacaga
     3121 acatccgcaa ggtcttcaag gacgtgcggg actcggtgct cgcccgctac ctggacgaga
     3181 tcaacctgct gtgatagctc gagtctagag ggcccgttta aacccgctga tcagcctcga
     3241 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     3301 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     3361 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     3421 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     3481 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     3541 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     3601 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     3661 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     3721 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag
     3781 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt
     3841 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat
     3901 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg
     3961 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga
     4021 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc
     4081 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc
     4141 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg
     4201 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga
     4261 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg
     4321 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc
     4381 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg
     4441 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct
     4501 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc
     4561 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac
     4621 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg
     4681 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg
     4741 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca
     4801 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct
     4861 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
     4921 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg
     4981 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     5041 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct
     5101 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     5161 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     5221 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     5281 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     5341 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5401 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5461 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5521 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     5581 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     5641 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     5701 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     5761 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     5821 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     5881 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     5941 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     6001 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     6061 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     6121 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     6181 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     6241 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     6301 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     6361 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     6421 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     6481 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     6541 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     6601 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     6661 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     6721 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     6781 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     6841 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     6901 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     6961 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     7021 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     7081 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     7141 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     7201 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     7261 cccgaaaagt gccacctgac gtc
//
