LOCUS       sequence-410281-        7064 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of REGA-SIGN, a NanoBRET-based G protein biosensor. It encodes 
            GNA12 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 7064)
  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 7064)
  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-12-133-NLuc
FEATURES             Location/Qualifiers
     source          1..7064
                     /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             1343..1852
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(2655..2672)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2661..2885
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2931..3359
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3018..3037)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3228..3249
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3368..3388)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3373..3702
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3553..3688
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3615..3634
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3769..4563
                     /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(3823..3842)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4433..4452
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4737..4858
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4774..4793)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4828..4847
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4907..4923)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4907..4923)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4920..4942)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4931..4947
                     /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(4955..4961,4962..4979,4980..4985))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5000..5021
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5138..5155)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5309..5897)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5389..5408)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6068..6859,6860..6928))
                     /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     6691..6710
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6929..7033)
                     /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 ccaccatgtc cggggtggtg cggaccctca gccgctgcct
      961 gctgccggcc gaggccggcg gggcccgcga gcgcagggcg ggcagcggcg cgcgcgacgc
     1021 ggagcgcgag gcccggaggc gtagccgcga catcgacgcg ctgctggccc gcgagcggcg
     1081 cgcggtccgg cgcctggtga agatcctgct gctgggcgcg ggcgagagcg gcaagtccac
     1141 gttcctcaag cagatgcgca tcatccacgg ccgcgagttc gaccagaagg cgctgctgga
     1201 gttccgcgac accatcttcg acaacatcct caagggctca agggttcttg ttgatgcacg
     1261 agataagctt ggcattcctt ggcagtattc tgaaaatgag aagcatggga tgttcctgat
     1321 ggccagcggc ggcggcggca gcgtcttcac actcgaagat ttcgttgggg actggcgaca
     1381 gacagccggc tacaacctgg accaagtcct tgaacaggga ggtgtgtcca gtttgtttca
     1441 gaatctcggg gtgtccgtaa ctccgatcca aaggattgtc ctgagcggtg aaaatgggct
     1501 gaagatcgac atccatgtca tcatcccgta tgaaggtctg agcggcgacc aaatgggcca
     1561 gatcgaaaaa atttttaagg tggtgtaccc tgtggatgat catcacttta aggtgatcct
     1621 gcactatggc acactggtaa tcgacggggt tacgccgaac atgatcgact atttcggacg
     1681 gccgtatgaa ggcatcgccg tgttcgacgg caaaaagatc actgtaacag ggaccctgtg
     1741 gaacggcaac aaaattatcg acgagcgcct gatcaacccc gacggctccc tgctgttccg
     1801 agtaaccatc aacggagtga ccggctggcg gctgtgcgaa cgcattctgg cgagcggcgg
     1861 cggcggcagc ttcgagaaca aggcggggct gcctgtggag ccggccacct tccagctgta
     1921 cgtcccggcc ctgagcgcac tctggaggga ttctggcatc agggaggctt tcagccggag
     1981 aagcgagttt cagctggggg agtcggtgaa gtacttcctg gacaacttgg accggatcgg
     2041 ccagctgaat tactttccta gtaagcaaga tatcctgctg gctaggaaag ccaccaaggg
     2101 aattgtggag catgacttcg ttattaagaa gatccccttt aagatggtgg atgtgggcgg
     2161 ccagcggtcc cagcgccaga agtggttcca gtgcttcgac gggatcacgt ccatcctgtt
     2221 catggtctcc tccagcgagt acgaccaggt cctcatggag gacaggcgca ccaaccggct
     2281 ggtggagtcc atgaacatct tcgagaccat cgtcaacaac aagctcttct tcaacgtctc
     2341 catcattctc ttcctcaaca agatggacct cctggtggag aaggtgaaga ccgtgagcat
     2401 caagaagcac ttcccggact tcaggggcga cccgcacagg ctggaggacg tccagcgcta
     2461 cctggtccag tgcttcgaca ggaagagacg gaaccgcagc aagccactct tccaccactt
     2521 caccaccgcc atcgacaccg agaacgtccg cttcgtgttc catgctgtga aagacaccat
     2581 cctgcaggag aacctgaagg acatcatgct gcagtgactc gagtctagag ggcccgttta
     2641 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     2701 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     2761 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     2821 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     2881 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     2941 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     3001 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     3061 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     3121 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     3181 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     3241 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     3301 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     3361 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3421 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3481 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3541 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3601 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     3661 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     3721 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     3781 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     3841 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     3901 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     3961 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     4021 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     4081 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     4141 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     4201 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     4261 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     4321 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     4381 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     4441 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     4501 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     4561 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     4621 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     4681 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     4741 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4801 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     4861 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     4921 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     4981 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     5041 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     5101 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     5161 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     5221 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     5281 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     5341 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     5401 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5461 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     5521 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5581 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5641 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5701 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     5761 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5821 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     5881 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5941 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     6001 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     6061 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     6121 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     6181 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     6241 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     6301 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     6361 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6421 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     6481 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6541 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6601 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6661 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6721 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6781 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6841 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6901 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     6961 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     7021 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtc
//
