LOCUS       sequence-301591-        6364 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a Gbeta subunit (GNB3) 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 6364)
  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 6364)
  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: pcDNA3.1-Beta3
FEATURES             Location/Qualifiers
     source          1..6364
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(66..654)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(146..165)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(825..1616,1617..1685))
                     /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     1448..1467
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1686..1790)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1865..1884)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2056..2435
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2436..2639
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2590..2610
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2684..2703
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2684..2702
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2716..2725
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3776..3793)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3782..4006
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4052..4480
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4139..4158)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4349..4370
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4489..4509)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4494..4823
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4674..4809
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4736..4755
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4890..5684
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4944..4963)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5554..5573
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5858..5979
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5895..5914)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5949..5968
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6028..6044)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6028..6044)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6041..6063)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6052..6068
                     /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(6076..6082,6083..6100,6101..6106))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6121..6142
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6259..6276)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
       61 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
      121 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
      181 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
      241 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
      301 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
      361 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
      421 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
      481 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
      541 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
      601 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
      661 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
      721 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
      781 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
      841 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
      901 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
      961 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     1021 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     1081 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     1141 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     1201 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     1261 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     1321 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     1381 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     1441 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     1501 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     1561 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     1621 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     1681 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     1741 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     1801 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
     1861 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt
     1921 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
     1981 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
     2041 ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca attacggggt
     2101 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
     2161 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
     2221 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
     2281 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
     2341 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
     2401 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
     2461 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
     2521 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
     2581 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctc
     2641 tctggctaac tagagaaccc actgcttact ggcttatcga aattaatacg actcactata
     2701 gggagaccca agcttgccac catgggggag atggagcagc tgaggcagga ggcggagcag
     2761 ctcaagaagc agattgcaga tgccaggaaa gcctgtgctg acgttactct ggcagagctg
     2821 gtgtctggcc tagaggtggt gggacgagtc cagatgcgga cgcggcggac gttaagggga
     2881 cacctggcca agatttacgc catgcactgg gccactgatt ctaagctgct ggtaagtgcc
     2941 tcgcaagatg ggaagctgat cgtgtgggac agctacacca ccaacaaggt gcacgccatc
     3001 ccactgcgct cctcctgggt catgacctgt gcctatgccc catcagggaa ctttgtggca
     3061 tgtggggggc tggacaacat gtgttccatc tacaacctca aatcccgtga gggcaatgtc
     3121 aaggtcagcc gggagctttc tgctcacaca ggttatctct cctgctgccg cttcctggat
     3181 gacaacaata ttgtgaccag ctcgggggac accacgtgtg ccttgtggga cattgagact
     3241 gggcagcaga agactgtatt tgtgggacac acgggtgact gcatgagcct ggctgtgtct
     3301 cctgacttca atctcttcat ttcgggggcc tgtgatgcca gtgccaagct ctgggatgtg
     3361 cgagagggga cctgccgtca gactttcact ggccacgagt cggacatcaa cgccatctgt
     3421 ttcttcccca atggagaggc catctgcacg ggctcggatg acgcttcctg ccgcttgttt
     3481 gacctgcggg cagaccagga gctgatctgc ttctcccacg agagcatcat ctgcggcatc
     3541 acgtccgtgg ccttctccct cagtggccgc ctactattcg ctggctacga cgacttcaac
     3601 tgcaatgtct gggactccat gaagtctgag cgtgtgggca tcctctctgg ccacgataac
     3661 agggtgagct gcctgggagt cacagctgac gggatggctg tggccacagg ttcctgggac
     3721 agcttcctca aaatctggaa ctgatctaga gggcccgttt aaacccgctg atcagcctcg
     3781 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     3841 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     3901 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     3961 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa
     4021 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg
     4081 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     4141 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     4201 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     4261 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     4321 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     4381 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     4441 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     4501 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     4561 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     4621 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     4681 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4741 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     4801 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     4861 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     4921 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     4981 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     5041 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg
     5101 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     5161 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     5221 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     5281 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     5341 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     5401 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     5461 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     5521 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     5581 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     5641 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     5701 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     5761 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     5821 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     5881 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     5941 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     6001 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     6061 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     6121 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     6181 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     6241 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     6301 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     6361 agga
//
