LOCUS       sequence-326675-        6157 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gaussia luciferase (GLucM23) targeted to the extracellular surface 
            of the cell membrane for bioluminescence resonance energy transfer 
            (BRET)-based measurement of ligand binding to unmodified receptors.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6157)
  AUTHORS   Toth AD, Garger D, Prokop S, Soltesz-Katona E, Varnai P, Balla A, 
            Turu G, Hunyady L
  TITLE     A general method for quantifying ligand binding to unmodified 
            receptors using Gaussia luciferase.
  JOURNAL   J Biol Chem. 2021 Feb 2:100366. doi: 10.1016/j.jbc.2021.100366.
  PUBMED    33545176
REFERENCE   2  (bases 1 to 6157)
  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: "J Biol 
            Chem. 2021 Feb 2:100366. doi: 10.1016/j.jbc.2021.100366."
COMMENT     Sequence Label: GLuc-PM
FEATURES             Location/Qualifiers
     source          1..6157
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      690..699
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             696..1250
                     /codon_start=1
                     /product="secreted Gaussia luciferase"
                     /label=hGLuc
                     /note="human codon-optimized"
                     /translation="MGVKVLFALICIAVAEAKPTENNEDFNIVAVASNFATTDLDADRG
                     KLPGEKLPLEVLKELEANARKAGCTRGCLICLSHIKCTPKMKKFIPGRCHTYEGDKESA
                     QGGIGEAIDDIPEIPGFKDLEPIEQFIAQVDLCVDCTTGCLKGLANVQCSDLLKKWLPQ
                     RCATFASKIQGQVDKIKGAGDD"
     CDS             1320..1466
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(1514..1531)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1520..1744
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1790..2218
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1877..1896)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2087..2108
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2227..2247)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2232..2561
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2412..2547
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2474..2493
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2628..3422
                     /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(2682..2701)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3292..3311
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3596..3717
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3633..3652)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3687..3706
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3766..3782)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3766..3782)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3779..3801)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3790..3806
                     /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(3814..3820,3821..3838,3839..3844))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3859..3880
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3997..4014)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4168..4756)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4248..4267)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4927..5718,5719..5787))
                     /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     5550..5569
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5788..5892)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5967..5986)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacttaagc ttaccggtcg ccaccatggg agtcaaagtt ctgtttgccc
      721 tgatctgcat cgctgtggcc gaggccaagc ccaccgagaa caacgaagac ttcaacatcg
      781 tggccgtggc cagcaacttc gcgaccacgg atctcgatgc tgaccgcggg aagttgcccg
      841 gcgagaagct gccgctggag gtgctcaaag agcttgaagc caatgcccgg aaagctggct
      901 gcaccagggg ctgtctgatc tgcctgtccc acatcaagtg cacgcccaag atgaagaagt
      961 tcatcccagg acgctgccac acctacgaag gcgacaaaga gtccgcacag ggcggcatag
     1021 gcgaggcgat cgatgacatt cctgagattc ctgggttcaa ggacttggag cccattgagc
     1081 agttcatcgc acaggtcgat ctgtgtgtgg actgcacaac tggctgcctc aaagggcttg
     1141 ccaacgtgca gtgttctgac ctgctcaaga agtggctgcc gcaacgctgt gcgacctttg
     1201 ccagcaagat ccagggccag gtggacaaga tcaagggggc cggtgatgac ggaggcggat
     1261 caggtggcgg atccagatct ggtggcggtt ctggtggcgg tagtggtggc ggtagtaatg
     1321 ctgtgggcca ggacacgcag gaggtcatcg tggtgccaca ctccttgccc tttaaggtgg
     1381 tggtgatctc agccatcctg gccctggtgg tgctcaccat catctccctt atcatcctca
     1441 tcatgctttg gcagaagaag ccacgttagg cggccgctcg agtctagagg gcccgtttaa
     1501 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     1561 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     1621 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     1681 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     1741 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     1801 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     1861 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     1921 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     1981 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     2041 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     2101 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     2161 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     2221 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2281 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     2341 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     2401 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     2461 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     2521 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     2581 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     2641 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     2701 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     2761 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     2821 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     2881 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     2941 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     3001 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     3061 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     3121 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     3181 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     3241 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     3301 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     3361 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     3421 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     3481 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     3541 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     3601 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     3661 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     3721 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     3781 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     3841 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     3901 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     3961 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4021 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4081 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4141 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4201 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4261 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4321 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4381 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4441 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     4501 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     4561 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     4621 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     4681 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     4741 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     4801 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     4861 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     4921 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     4981 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5041 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5101 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5161 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5221 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5281 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5341 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5401 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     5461 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     5521 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     5581 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     5641 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     5701 aaccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     5761 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     5821 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     5881 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct
     5941 cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     6001 tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct
     6061 acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt
     6121 gcgctgcttc gcgatgtacg ggccagatat acgcgtt
//
