LOCUS       sequence-401597-        5995 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GPCR/G protein BRET biosensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5995)
  AUTHORS   Janicot R, Maziarz M, Park JC, Zhao J, Luebbers A, Green E, 
            Philibert CE, Zhang H, Layne MD, Wu JC, Garcia-Marcos M
  TITLE     Direct interrogation of context-dependent GPCR activity with a 
            universal biosensor platform.
  JOURNAL   Cell. 2024 Feb 16:S0092-8674(24)00065-5. doi: 
            10.1016/j.cell.2024.01.028.
  PUBMED    38412860
REFERENCE   2  (bases 1 to 5995)
  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: "Cell. 2024 
            Feb 16:S0092-8674(24)00065-5. doi: 10.1016/j.cell.2024.01.028."
COMMENT     Sequence Label: pcDNA3.1-mas-KB2123-Nluc
FEATURES             Location/Qualifiers
     source          1..5995
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(103..122)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        294..673
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        674..877
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     828..848
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     922..941
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        922..940
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             963..983
                     /codon_start=1
                     /product="N-myristoylation signal from Src kinase (Pellman 
                     et al., 1985; Kaplan et al., 1988)"
                     /label=myr
                     /translation="MGSSKSK"
     CDS             1092..1607
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     primer_bind     complement(1645..1662)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1651..1875
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1921..2349
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2008..2027)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2218..2239
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2358..2378)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2363..2692
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2543..2678
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2605..2624
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2759..3553
                     /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(2813..2832)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3423..3442
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3727..3848
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3764..3783)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3818..3837
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3897..3913)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3897..3913)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3910..3932)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3921..3937
                     /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(3945..3951,3952..3969,3970..3975))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3990..4011
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4128..4145)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4299..4887)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4379..4398)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5058..5849,5850..5918))
                     /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     5681..5700
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5919..5995,1..28))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
       61 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
      121 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
      181 gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct
      241 tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg
      301 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
      361 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
      421 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      481 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      541 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      601 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      661 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
      721 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
      781 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
      841 taggcgtgta cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac
      901 tgcttactgg cttatcgaaa ttaatacgac tcactatagg gagacccaag ctggctagca
      961 ccatggggag tagcaagagc aagacgtcga attccggaat caagcttggt ggatcatcag
     1021 gaggtagcag cagcgagctg gccgactact tcggctggga cggctggccc ggcagcggag
     1081 gtggtggatc catggtcttc acactcgaag atttcgttgg ggactggcga cagacagccg
     1141 gctacaacct ggaccaagtc cttgaacagg gaggtgtgtc cagtttgttt cagaatctcg
     1201 gggtgtccgt aactccgatc caaaggattg tcctgagcgg tgaaaatggg ctgaagatcg
     1261 acatccatgt catcatcccg tatgaaggtc tgagcggcga ccaaatgggc cagatcgaaa
     1321 aaatttttaa ggtggtgtac cctgtggatg atcatcactt taaggtgatc ctgcactatg
     1381 gcacactggt aatcgacggg gttacgccga acatgatcga ctatttcgga cggccgtatg
     1441 aaggcatcgc cgtgttcgac ggcaaaaaga tcactgtaac agggaccctg tggaacggca
     1501 acaaaattat cgacgagcgc ctgatcaacc ccgacggctc cctgctgttc cgagtaacca
     1561 tcaacggagt gaccggctgg cggctgtgcg aacgcattct ggcgtaactc gagtctagag
     1621 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     1681 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     1741 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     1801 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     1861 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     1921 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     1981 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     2041 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     2101 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     2161 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     2221 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     2281 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     2341 acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     2401 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     2461 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     2521 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     2581 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga
     2641 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc
     2701 gggagcttgt atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat
     2761 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     2821 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     2881 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     2941 ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     3001 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     3061 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     3121 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     3181 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     3241 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg
     3301 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     3361 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     3421 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     3481 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     3541 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     3601 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     3661 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     3721 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     3781 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     3841 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     3901 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     3961 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     4021 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     4081 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     4141 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4201 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4261 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4321 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4381 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4441 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4501 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4561 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4621 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4681 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     4741 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4801 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4861 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     4921 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     4981 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5041 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5101 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5161 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5221 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     5281 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     5341 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     5401 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     5461 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5521 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5581 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     5641 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     5701 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     5761 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     5821 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     5881 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     5941 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tattt
//
