LOCUS       sequence-301587-        7292 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a G alpha subunit (GNA13) 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 7292)
  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 7292)
  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-GAlpha13-RLuc8
FEATURES             Location/Qualifiers
     source          1..7292
                     /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             1494..2426
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNATSSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGAALAFHYAYEHQDRIKAIVHMESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETVLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKLFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFLQEDAPDEMGKYIKSFVERVLKNEQ"
     primer_bind     complement(3244..3261)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3250..3474
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3607..3626)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3758..3805
                     /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             3813..4835
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3813..4835
                     /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    4965..5086
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5002..5021)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5056..5075
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5135..5151)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5135..5151)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5148..5170)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5159..5175
                     /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(5183..5189,5190..5207,5208..5213))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5228..5249
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5366..5383)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5537..6125)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5617..5636)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6296..7087,7088..7156))
                     /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     6919..6938
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7157..7261)
                     /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 atggcggact tcctgccgtc gcggtccgtg ctgtccgtgt
     1141 gcttccccgg ctgcctgctg acgagtggcg aggccgagca gcaacgcaag tccaaggaga
     1201 tcgacaaatg cctgtctcgg gaaaagacct atgtgaagcg gctggtgaag atcctgctgc
     1261 tgggcgcggg cgagagcggc aagtccacct tcctgaagca gatgcggatc atccacgggc
     1321 aggacttcga ccagcgcgcg cgcgaggagt tccgccccac catctacagc aacgtgatca
     1381 aaggtatgag ggtgctggtt gatgctcgag agaagcttca tattccctgg ggagacaact
     1441 caaaccaaca acatggagat aagatgatgt cgttttctgg tggtggtgga tccatggctt
     1501 ccaaggtgta cgaccccgag caacgcaaac gcatgatcac tgggcctcag tggtgggctc
     1561 gctgcaagca aatgaacgtg ctggactcct tcatcaacta ctatgattcc gagaagcacg
     1621 ccgagaacgc cgtgattttt ctgcatggta acgctacctc cagctacctg tggaggcacg
     1681 tcgtgcctca catcgagccc gtggctagat gcatcatccc tgatctgatc ggaatgggta
     1741 agtccggcaa gagcgggaat ggctcatatc gcctcctgga tcactacaag tacctcaccg
     1801 cttggttcga gctgctgaac cttccaaaga aaatcatctt tgtgggccac gactgggggg
     1861 ctgctctggc ctttcactac gcctacgagc accaagacag gatcaaggcc atcgtccata
     1921 tggagagtgt cgtggacgtg atcgagtcct gggacgagtg gcctgacatc gaggaggata
     1981 tcgccctgat caagagcgaa gagggcgaga aaatggtgct tgagaataac ttcttcgtcg
     2041 agaccgtgct cccaagcaag atcatgcgga aactggagcc tgaggagttc gctgcctacc
     2101 tggagccatt caaggagaag ggcgaggtta gacggcctac cctctcctgg cctcgcgaga
     2161 tccctctcgt taagggaggc aagcccgacg tcgtccagat tgtccgcaac tacaacgcct
     2221 accttcgggc cagcgacgat ctgcctaagc tgttcatcga gtccgaccct gggttctttt
     2281 ccaacgctat tgtcgaggga gctaagaagt tccctaacac cgagttcgtg aaggtgaagg
     2341 gcctccactt cctccaggag gacgctccag atgaaatggg taagtacatc aagagcttcg
     2401 tggagcgcgt gctgaagaac gagcagagcg gaggaggcgg cagtgatacc cgggccccca
     2461 tggcagccca aggaatggtg gaaacaaggg ttttcttaca atatcttcct gctataagag
     2521 cattatgggc agacagcggc atacagaatg cctatgaccg gcgtcgagaa tttcaactgg
     2581 gtgaatctgt aaaatatttc ctggataact tggataaact tggagaacca gattatattc
     2641 catcacaaca agatattctg cttgccagaa gacccaccaa aggcatccat gaatacgact
     2701 ttgaaataaa aaatgttcct ttcaaaatgg ttgatgtagg tggtcagaga tcagaaagga
     2761 aacgttggtt tgaatgtttc gacagtgtga catcaatact tttccttgtt tcctcaagtg
     2821 aatttgacca ggtgcttatg gaagatcgac tgaccaatcg ccttacagag tctctgaaca
     2881 tttttgaaac aatcgtcaat aaccgggttt tcagcaatgt ctccataatt ctgttcttaa
     2941 acaagacaga cttgcttgag gagaaggtgc aaattgtgag catcaaagac tatttcctag
     3001 aatttgaagg ggatccccac tgcttaagag acgtccaaaa attcctggtg gaatgtttcc
     3061 ggaacaaacg ccgggaccag caacagaagc ccttatacca ccacttcacc actgctatca
     3121 acacggagaa catccgcctt gttttccgtg acgtgaagga tactattctg catgacaacc
     3181 tcaagcagct tatgctacag tgatagctcg agtctagagg gcccgtttaa acccgctgat
     3241 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
     3301 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
     3361 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
     3421 gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg
     3481 aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat
     3541 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     3601 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     3661 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     3721 ccaaaaaact tgattagggt gatggttcac gtacctagaa gttcctattc cgaagttcct
     3781 attctctaga aagtatagga acttccttgg ccaaaaagcc tgaactcacc gcgacgtctg
     3841 tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg
     3901 gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa
     3961 atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg
     4021 cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca
     4081 tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg
     4141 ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga
     4201 gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca
     4261 tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca
     4321 gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag
     4381 tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca
     4441 taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca
     4501 acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc
     4561 ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc
     4621 ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg
     4681 gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc
     4741 gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc
     4801 gacgccccag cactcgtccg agggcaaagg aatagcacgt actacgagat ttcgattcca
     4861 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
     4921 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
     4981 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     5041 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
     5101 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     5161 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     5221 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     5281 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     5341 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     5401 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     5461 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5521 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5581 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5641 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     5701 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     5761 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     5821 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     5881 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     5941 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     6001 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6061 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6121 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6181 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6241 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6301 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6361 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     6421 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     6481 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     6541 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     6601 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6661 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     6721 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     6781 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     6841 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     6901 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     6961 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     7021 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     7081 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     7141 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     7201 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     7261 gcacatttcc ccgaaaagtg ccacctgacg tc
//
