LOCUS       sequence-410279-        6977 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of REGA-SIGN, a NanoBRET-based G protein biosensor. It encodes 
            transcript variant B of GNAO tagged with NanoLuciferase and is meant
            to be paired with Ggamma2 labeled with LSS-mKATE2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6977)
  AUTHORS   Boon K, Vanalken N, Meyen E, Schols D, Van Loy T
  TITLE     REGA-SIGN: Development of a Novel Set of NanoBRET-Based G Protein 
            Biosensors.
  JOURNAL   Biosensors (Basel). 2023 Jul 28;13(8):767. doi: 
            10.3390/bios13080767.
  PUBMED    37622853
REFERENCE   2  (bases 1 to 6977)
  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: "Biosensors 
            (Basel)."; date: "2023-07-28"; volume: "13(8):767. doi"; pages: " 
            10.3390/bios13080767"
COMMENT     Sequence Label: pcDNA3.1(+)G-alpha-oB-91-NLuc
FEATURES             Location/Qualifiers
     source          1..6977
                     /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             1217..1726
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(2568..2585)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2574..2798
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2844..3272
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2931..2950)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3141..3162
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3281..3301)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3286..3615
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3466..3601
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3528..3547
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3682..4476
                     /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(3736..3755)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4346..4365
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4650..4771
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4687..4706)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4741..4760
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4820..4836)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4820..4836)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4833..4855)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4844..4860
                     /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(4868..4874,4875..4892,4893..4898))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4913..4934
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5051..5068)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5222..5810)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5302..5321)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5981..6772,6773..6841))
                     /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     6604..6623
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6842..6946)
                     /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 gtttaaactt aagcttggta ccaccatggg atgtactctg agcgcagagg agagagccgc
      961 cctcgagcgg agcaaggcga ttgagaaaaa cctcaaagag gatggcatca gcgccgccaa
     1021 agacgtgaaa ttactcctgc tcggggctgg agaatcagga aaaagcacca ttgtgaagca
     1081 gatgaagatc atccatgaag atggcttctc cggagaagac gtgaaacagt acaagcctgt
     1141 tgtctacagc aacactatcc agtccctggc agccatcgtc cgggccatgg acactttgag
     1201 cggcggcggc ggcagcgtct tcacactcga agatttcgtt ggggactggc gacagacagc
     1261 cggctacaac ctggaccaag tccttgaaca gggaggtgtg tccagtttgt ttcagaatct
     1321 cggggtgtcc gtaactccga tccaaaggat tgtcctgagc ggtgaaaatg ggctgaagat
     1381 cgacatccat gtcatcatcc cgtatgaagg tctgagcggc gaccaaatgg gccagatcga
     1441 aaaaattttt aaggtggtgt accctgtgga tgatcatcac tttaaggtga tcctgcacta
     1501 tggcacactg gtaatcgacg gggttacgcc gaacatgatc gactatttcg gacggccgta
     1561 tgaaggcatc gccgtgttcg acggcaaaaa gatcactgta acagggaccc tgtggaacgg
     1621 caacaaaatt atcgacgagc gcctgatcaa ccccgacggc tccctgctgt tccgagtaac
     1681 catcaacgga gtgaccggct ggcggctgtg cgaacgcatt ctggcgagcg gcggcggcgg
     1741 cagcggcatc gaatatggtg ataaggagag aaaggctgac gccaagatgg tgtgtgatgt
     1801 ggtgagtcgg atggaagaca ccgagccctt ctctgcagag ctgctttctg ccatgatgcg
     1861 gctctggggc gactcaggaa tccaagagtg cttcaaccgg tcccgggagt atcagctcaa
     1921 cgactctgcc aaatactacc tggacagcct ggatcggatt ggggccgccg actaccagcc
     1981 caccgagcag gacatcctcc gaaccagggt caaaaccact ggcatcgtag aaacccactt
     2041 cacattcaag aacctccact tcaggctgtt tgacgtcgga ggccagcgat ctgaacgcaa
     2101 gaagtggatc cattgcttcg aggacgtcac ggccatcatt ttctgtgtcg cgctcagcgg
     2161 ctatgaccag gtgctccacg aagacgaaac cacgaaccgc atgcacgaat ccctgaagct
     2221 ttttgacagc atctgcaaca acaaatggtt cacagacacg tccatcatcc tgtttcttaa
     2281 caagaaggac atatttgaag agaagatcaa gaagtccccg ctcaccatct gctttcctga
     2341 atatacaggc cccagcgcct tcacagaagc cgtggcttac atccaggccc agtacgagag
     2401 caagaacaag tcagcccaca aggagatcta cacccacgtc acctgcgcca cggacaccaa
     2461 caacatccag tttgtctttg atgctgtgac ggacgtcatc atcgccaaaa acctgcgggg
     2521 ctgtggactc tactgatcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc
     2581 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     2641 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     2701 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     2761 caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag
     2821 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     2881 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     2941 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     3001 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     3061 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     3121 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     3181 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     3241 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     3301 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3361 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3421 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3481 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3541 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3601 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag
     3661 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     3721 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     3781 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     3841 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     3901 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     3961 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     4021 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     4081 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     4141 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     4201 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     4261 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     4321 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     4381 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     4441 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     4501 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     4561 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     4621 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     4681 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     4741 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     4801 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     4861 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     4921 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     4981 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     5041 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     5101 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     5161 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     5221 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     5281 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     5341 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     5401 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     5461 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     5521 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     5581 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     5641 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     5701 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     5761 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     5821 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     5881 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     5941 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6001 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     6061 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     6121 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     6181 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     6241 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     6301 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6361 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6421 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6481 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6541 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6601 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6661 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6721 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6781 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6841 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6901 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6961 aagtgccacc tgacgtc
//
