LOCUS       sequence-410283-        7097 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of REGA-SIGN, a NanoBRET-based G protein biosensor. It encodes 
            transcript variant A 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 7097)
  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 7097)
  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-oA-91-NLuc
FEATURES             Location/Qualifiers
     source          1..7097
                     /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             1276..1785
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(2688..2705)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2694..2918
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2964..3392
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3051..3070)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3261..3282
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3401..3421)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3406..3735
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3586..3721
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3648..3667
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3802..4596
                     /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(3856..3875)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4466..4485
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4770..4891
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4807..4826)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4861..4880
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4940..4956)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4940..4956)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4953..4975)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4964..4980
                     /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(4988..4994,4995..5012,5013..5018))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5033..5054
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5171..5188)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5342..5930)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5422..5441)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6101..6892,6893..6961))
                     /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     6724..6743
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6962..7066)
                     /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 ccgagctcgg atccactagt aacggccgcc agtgtgctgg
      961 aattcgccct tgatcggtac caccatggga tgtactctga gcgcagagga gagagccgcc
     1021 ctcgagcgga gcaaggcgat tgagaaaaac ctcaaagagg atggcatcag cgccgccaaa
     1081 gacgtgaaat tactcctgct cggggctgga gaatcaggaa aaagcaccat tgtgaagcag
     1141 atgaagatca tccatgaaga tggcttctcc ggagaagacg tgaaacagta caagcctgtt
     1201 gtctacagca acactatcca gtccctggca gccatcgtcc gggccatgga cactttgagc
     1261 ggcggcggcg gcagcgtctt cacactcgaa gatttcgttg gggactggcg acagacagcc
     1321 ggctacaacc tggaccaagt ccttgaacag ggaggtgtgt ccagtttgtt tcagaatctc
     1381 ggggtgtccg taactccgat ccaaaggatt gtcctgagcg gtgaaaatgg gctgaagatc
     1441 gacatccatg tcatcatccc gtatgaaggt ctgagcggcg accaaatggg ccagatcgaa
     1501 aaaattttta aggtggtgta ccctgtggat gatcatcact ttaaggtgat cctgcactat
     1561 ggcacactgg taatcgacgg ggttacgccg aacatgatcg actatttcgg acggccgtat
     1621 gaaggcatcg ccgtgttcga cggcaaaaag atcactgtaa cagggaccct gtggaacggc
     1681 aacaaaatta tcgacgagcg cctgatcaac cccgacggct ccctgctgtt ccgagtaacc
     1741 atcaacggag tgaccggctg gcggctgtgc gaacgcattc tggcgagcgg cggcggcggc
     1801 agcggcatcg aatatggtga taaggagaga aaggctgacg ccaagatggt gtgtgatgtg
     1861 gtgagtcgga tggaagacac cgagcccttc tctgcagagc tgctttctgc catgatgcgg
     1921 ctctggggcg actcaggaat ccaagagtgc ttcaaccggt cccgggagta tcagctcaac
     1981 gactctgcca aatactacct ggacagcctg gatcggattg gggccgccga ctaccagccc
     2041 accgagcagg acatcctccg aaccagggtc aaaaccactg gcatcgtaga aacccacttc
     2101 acattcaaga acctccactt caggctgttt gacgtcggag gccagcgatc tgaacgcaag
     2161 aagtggatcc attgcttcga ggacgtcacg gccatcattt tctgtgtcgc gctcagcggc
     2221 tatgaccagg tgctccacga agacgaaacc acgaaccgca tgcacgagtc tctcatgctc
     2281 ttcgactcca tctgtaacaa caagttcttc atcgatacct ccatcattct cttcctcaac
     2341 aagaaagatc tctttggcga gaagatcaag aagtcacctt tgaccatctg ctttcctgaa
     2401 tacacaggcc ccaataccta tgaagacgca gccgcctaca tccaagcaca atttgaaagc
     2461 aaaaaccgct cacccaacaa agaaatatat tgtcacatga cttgtgccac agacacgaat
     2521 aacatccagg tggtgttcga cgccgtcacc gacatcatca ttgccaacaa cctccggggc
     2581 tgcggcttgt actgatctag agatcaaggg cgaattctgc agatatccat cacactggcg
     2641 gccgctcgag catgcatcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc
     2701 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     2761 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     2821 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     2881 caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag
     2941 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     3001 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     3061 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     3121 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     3181 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     3241 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     3301 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     3361 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg
     3421 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3481 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3541 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3601 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3661 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3721 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag
     3781 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc
     3841 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     3901 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     3961 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     4021 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     4081 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     4141 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     4201 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     4261 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     4321 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     4381 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     4441 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     4501 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     4561 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga
     4621 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg
     4681 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
     4741 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
     4801 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     4861 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
     4921 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     4981 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     5041 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     5101 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     5161 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     5221 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     5281 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     5341 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     5401 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     5461 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     5521 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     5581 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     5641 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     5701 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     5761 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     5821 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     5881 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     5941 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     6001 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     6061 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6121 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     6181 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     6241 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     6301 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     6361 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     6421 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     6481 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     6541 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     6601 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     6661 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     6721 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     6781 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     6841 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     6901 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     6961 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     7021 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     7081 aagtgccacc tgacgtc
//
