LOCUS       sequence-387096-        7481 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human FZD8 with an N-terminal HiBiT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7481)
  AUTHORS   Kozielewicz P, Shekhani R, Moser S, Bowin CF, Wesslowski J, Davidson
            G, Schulte G
  TITLE     Quantitative Profiling of WNT-3A Binding to All Human Frizzled 
            Paralogues in HEK293 Cells by NanoBiT/BRET Assessments.
  JOURNAL   ACS Pharmacol Transl Sci. 2021 May 11;4(3):1235-1245. doi: 
            10.1021/acsptsci.1c00084. eCollection 2021 Jun 11.
  PUBMED    34151213
REFERENCE   2  (bases 1 to 7481)
  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: 
            "10.1021/acsptsci.1c00084"; journalName: "ACS Pharmacol Transl Sci";
            date: "2021-05-11- 11"; volume: "4"; issue: "3"; pages: "1235-1245"
COMMENT     Sequence Label: HiBiT-FZD8
FEATURES             Location/Qualifiers
     source          1..7481
                     /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"
     primer_bind     complement(3072..3089)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3078..3302
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3348..3776
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3435..3454)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3645..3666
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3785..3805)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3790..4119
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3970..4105
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4032..4051
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4186..4980
                     /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(4240..4259)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4850..4869
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5154..5275
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5191..5210)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5245..5264
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5324..5340)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5324..5340)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5337..5359)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5348..5364
                     /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(5372..5378,5379..5396,5397..5402))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5417..5438
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5555..5572)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5726..6314)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5806..5825)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6485..7276,7277..7345))
                     /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     7108..7127
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7346..7450)
                     /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 aagcttgcca ccatgcggct ctgcatcccg caggtgctgt tggccttgtt
      961 cctttccatg ctgacagggc cgggagaagg cagccgggtg agcggctggc ggctgttcaa
     1021 gaagattagc ggatccgcga gtgcgaagga acttgcatgt caggagatca cggttccact
     1081 gtgcaaaggt atagggtaca actatactta catgcctaat caatttaacc atgacaccca
     1141 agatgaagcg ggcctggagg tccaccaatt ctggccgctt gtggaaatcc aatgtagccc
     1201 ggatcttaag tttttcttgt gttccatgta tacgccaatt tgcctggagg attataaaaa
     1261 gcccctcccg ccttgccgct cagtttgtga acgggcgaag gctgggtgcg caccccttat
     1321 gaggcaatat ggcttcgcgt ggccagatcg catgagatgt gatcgcttgc cggaacaagg
     1381 gaatcctgat actctgtgta tggactataa caggacggat ctcaccaccg cggcaccgtc
     1441 ccctccccgg aggttgcctc ctccaccacc gggcgagcag cccccaagtg gaagtggcca
     1501 tgggagaccg ccgggagcta ggcctccaca tcgcggaggt ggaaggggtg gtggaggcgg
     1561 tgatgcggct gcaccgcctg ctcgcggggg cggtggcggg ggaaaagcta gaccaccggg
     1621 cggcggtgcg gccccttgtg agccgggttg tcagtgccgc gctcccatgg tttccgtaag
     1681 tagtgaacgc cacccgttgt ataatcgggt gaaaaccgga caaatagcaa attgcgccct
     1741 gccctgtcat aacccctttt tctcccagga cgaacgcgcg tttacagtct tctggattgg
     1801 tctttggtcc gttctttgtt ttgtatctac ttttgctacc gttagtacat ttttgatcga
     1861 catggaacgg ttcaagtacc ctgagcggcc tattattttt ctctccgctt gctacctgtt
     1921 cgtgtccgtt gggtacctgg ttagactggt agcggggcac gagaaagtcg cttgttcagg
     1981 tggagcacct ggagcaggag gagcaggagg agctggaggg gccgccgcgg gtgcgggcgc
     2041 tgctggcgct ggggcaggtg gacctggggg tagaggcgag tacgaagaac ttggagcagt
     2101 ggagcaacat gtacgctacg agaccacagg tccagctttg tgtacagttg tatttttgct
     2161 ggtatacttc tttggaatgg ctagctctat ctggtgggtc atcctttccc tcacatggtt
     2221 ccttgctgcg ggtatgaaat ggggcaatga ggcgatcgca ggatacagtc aatacttcca
     2281 tcttgcggca tggcttgtac cctctgtaaa gtccattgca gttctggcac tcagctctgt
     2341 tgatggagac cctgtcgcag gtatatgcta tgtcggaaat cagtccctgg ataatcttag
     2401 ggggtttgtg cttgcgcctt tggtgattta tctctttatt ggtaccatgt tcctccttgc
     2461 tggcttcgtt agtttgtttc ggatcaggag tgtaataaag cagcaggatg gcccaactaa
     2521 gacccataaa ctcgaaaaac ttatgatccg cttgggactt ttcaccgtcc tctacaccgt
     2581 accggctgcg gtcgttgttg catgcctttt ttacgaacaa cataacaggc cgagatggga
     2641 ggccacacat aattgcccat gtcttcgaga tctgcaacca gatcaggcac gaagaccgga
     2701 ttatgctgtc tttatgttga aatacttcat gtgcctcgtt gtaggtatta catccggcgt
     2761 gtgggtttgg agcggaaaaa cactggaatc atggcgctcc ttgtgcacta gatgctgctg
     2821 ggcgagtaaa ggagcagcag taggaggagg tgctggagcc actgcggctg gaggaggagg
     2881 aggacctgga ggaggaggtg gcggcgggcc tggcggcggt ggaggccccg gtggaggggg
     2941 gggcagtctc tattccgacg tttctactgg tttgacgtgg cgatccggaa cagcttcatc
     3001 tgtttcctat cctaaacaga tgccattgag tcaggtttaa tctagagggc ccgtttaaac
     3061 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
     3121 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
     3181 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
     3241 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     3301 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     3361 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     3421 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     3481 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     3541 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     3601 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     3661 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     3721 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     3781 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     3841 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     3901 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     3961 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     4021 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     4081 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     4141 tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     4201 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     4261 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     4321 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
     4381 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     4441 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     4501 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     4561 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     4621 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     4681 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc
     4741 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     4801 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     4861 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     4921 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     4981 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     5041 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     5101 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt
     5161 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     5221 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     5281 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     5341 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     5401 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     5461 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     5521 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     5581 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     5641 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     5701 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     5761 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     5821 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     5881 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     5941 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     6001 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     6061 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     6121 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     6181 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     6241 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     6301 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     6361 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     6421 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     6481 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     6541 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     6601 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     6661 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     6721 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     6781 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     6841 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     6901 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     6961 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     7021 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     7081 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     7141 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     7201 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     7261 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     7321 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     7381 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     7441 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
