LOCUS       sequence-306384-        7051 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nanoluc complementation assay. Expression of G-alpha-12 protein 
            fused with the LgBiT(LgB)of the nanoluciferase inserted between 
            residues 115 and 116 of G-alpha-12.Addition of the HA epitope at N 
            terminus of G-alpha-12..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7051)
  AUTHORS   Laschet C, Dupuis N, Hanson J
  TITLE     A dynamic and screening-compatible nanoluciferase-based 
            complementation assay enables profiling of individual GPCR-G protein
            interactions.
  JOURNAL   J Biol Chem. 2019 Mar 15;294(11):4079-4090. doi: 
            10.1074/jbc.RA118.006231. Epub 2018 Dec 28.
  PUBMED    30593506
REFERENCE   2  (bases 1 to 7051)
  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"; doi: 
            "10.1074/jbc.RA118.006231"; journalName: "J Biol Chem"; date: 
            "2019-03-15- 15"; volume: "294"; issue: "11"; pages: "4079-4090"
COMMENT     Sequence Label: pcDNA3.1.G-alpha-12-LgB115
FEATURES             Location/Qualifiers
     source          1..7051
                     /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"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             907..933
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2642..2659)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2648..2872
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2918..3346
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3005..3024)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3215..3236
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3355..3375)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3360..3689
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3540..3675
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3602..3621
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3756..4550
                     /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(3810..3829)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4420..4439
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4724..4845
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4761..4780)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4815..4834
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4894..4910)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4894..4910)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4907..4929)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4918..4934
                     /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(4942..4948,4949..4966,4967..4972))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4987..5008
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5125..5142)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5296..5884)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5376..5395)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6055..6846,6847..6915))
                     /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     6678..6697
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6916..7020)
                     /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 ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 accatgtacc catacgatgt tccagattac gcttccgggg tggtgcggac cctcagccgc
      961 tgcctgctgc cggccgaggc cggcggggcc cgcgagcgca gggcgggcag cggcgcgcgc
     1021 gacgcggagc gcgaggcccg gaggcgtagc cgcgacatcg acgcgctgct ggcccgcgag
     1081 cggcgcgcgg tccggcgcct ggtgaagatc ctgctgctgg gcgcgggcga gagcggcaag
     1141 tccacgttcc tcaagcagat gcgcatcatc cacggccgcg agttcgacca gaaggcgctg
     1201 ctggagttcc gcgacaccat cttcgacaac atcctcaagg gctcaagggt tcttgttgat
     1261 gcacgagata agcttggatc cagcggaggc ggaggaagcg tcttcacact cgaagatttc
     1321 gttggggact gggaacagac agccgcctac aacctggacc aagtccttga acagggaggt
     1381 gtgtccagtt tgctgcagaa tctcgccgtg tccgtaactc cgatccaaag gattgtccgg
     1441 agcggtgaaa atgccctgaa gatcgacatc catgtcatca tcccgtatga aggtctgagc
     1501 gccgaccaaa tggcccagat cgaagaggtg tttaaggtgg tgtaccctgt ggatgatcat
     1561 cactttaagg tgatcctgcc ctatggcaca ctggtaatcg acggggttac gccgaacatg
     1621 ctgaactatt tcggacggcc gtatgaaggc atcgccgtgt tcgacggcaa aaagatcact
     1681 gtaacaggga ccctgtggaa cggcaacaaa attatcgacg agcgcctgat cacccccgac
     1741 ggctccatgc tgttccgagt aaccatcaac agtagcggag gcggaggaag cccgcggggc
     1801 attccttggc agtattctga aaatgagaag catgggatgt tcctgatggc cttcgagaac
     1861 aaggcggggc tgcctgtgga gccggccacc ttccagctgt acgtcccggc cctgagcgca
     1921 ctctggaggg attctggcat cagggaggct ttcagccgga gaagcgagtt tcagctgggg
     1981 gagtcggtga agtacttcct ggacaacttg gaccggatcg gccagctgaa ttactttcct
     2041 agtaagcaag atatcctgct ggctaggaaa gccaccaagg gaattgtgga gcatgacttc
     2101 gttattaaga agatcccctt taagatggtg gatgtgggcg gccagcggtc ccagcgccag
     2161 aagtggttcc agtgcttcga cgggatcacg tccatcctgt tcatggtctc ctccagcgag
     2221 tacgaccagg tcctcatgga ggacaggcgc accaaccggc tggtggagtc catgaacatc
     2281 ttcgagacca tcgtcaacaa caagctcttc ttcaacgtct ccatcattct cttcctcaac
     2341 aagatggacc tcctggtgga gaaggtgaag accgtgagca tcaagaagca cttcccggac
     2401 ttcaggggcg acccgcacag gctggaggac gtccagcgct acctggtcca gtgcttcgac
     2461 aggaagagac ggaaccgcag caagccactc ttccaccact tcaccaccgc catcgacacc
     2521 gagaacgtcc gcttcgtgtt ccatgctgtg aaagacacca tcctgcagga gaacctgaag
     2581 gacatcatgc tgcagtgaga attcctcgag tctagagggc ccgtttaaac ccgctgatca
     2641 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     2701 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
     2761 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
     2821 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag
     2881 gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta
     2941 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     3001 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     3061 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     3121 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     3181 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     3241 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     3301 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg
     3361 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3421 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3481 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
     3541 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     3601 ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag
     3661 gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg
     3721 gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     3781 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     3841 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     3901 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
     3961 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     4021 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     4081 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     4141 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     4201 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     4261 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
     4321 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     4381 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     4441 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     4501 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     4561 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     4621 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     4681 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     4741 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4801 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     4861 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     4921 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4981 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     5041 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     5101 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     5161 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     5221 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     5281 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     5341 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     5401 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5461 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     5521 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5581 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5641 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5701 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5761 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5821 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5881 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5941 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     6001 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     6061 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     6121 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     6181 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     6241 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     6301 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     6361 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6421 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6481 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6541 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6601 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6661 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6721 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6781 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6841 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6901 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6961 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     7021 cacatttccc cgaaaagtgc cacctgacgt c
//
