LOCUS       sequence-209698-        6524 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of a chimeric G-protein coupled receptor 
            (Rhodopsin-GPR174; Opto-GPR174; F1).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6524)
  AUTHORS   Morri M, Sanchez-Romero I, Tichy AM, Kainrath S, Gerrard EJ, 
            Hirschfeld PP, Schwarz J, Janovjak H
  TITLE     Optical functionalization of human Class A orphan G-protein-coupled 
            receptors.
  JOURNAL   Nat Commun. 2018 May 16;9(1):1950. doi: 10.1038/s41467-018-04342-1.
  PUBMED    29769519
REFERENCE   2  (bases 1 to 6524)
  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: "Nat 
            Commun."; date: "2018-05-16"; volume: "9(1):1950. doi"; pages: " 
            10.1038/s41467-018-04342-1"
COMMENT     Sequence Label: Opto-GPR174
FEATURES             Location/Qualifiers
     source          1..6524
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        74..453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        454..657
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     608..628
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     702..721
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        702..720
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             813..845
                     /codon_start=1
                     /product="epitope tag from vesicular stomatitis virus G 
                     protein"
                     /label=VSV-G tag
                     /translation="YTDIEMNRLGK"
     CDS             870..908
                     /codon_start=1
                     /product="segment of bovine rhodopsin containing a signal 
                     for N-linked glycosylation (Favaloro et al., 2008)"
                     /label=opsin tag
                     /note="The asparagine in the NKT tripeptide will be 
                     glycosylated if this sequence is exposed to the ER lumen."
                     /translation="GPNFYVPFSNKTG"
     primer_bind     complement(1954..1971)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1960..2184
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2230..2658
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2317..2336)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2527..2548
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2667..2687)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2672..3001
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2852..2987
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2914..2933
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3068..3862
                     /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(3122..3141)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3732..3751
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4036..4157
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4073..4092)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4127..4146
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4206..4222)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4206..4222)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4219..4241)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4230..4246
                     /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(4254..4260,4261..4278,4279..4284))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4299..4320
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4437..4454)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4608..5196)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4688..4707)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5367..6158,6159..6227))
                     /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     5990..6009
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6228..6332)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6407..6426)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
       61 agatatacgc gttgacattg attattgact agttattaat agtaatcaat tacggggtca
      121 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      181 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      241 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      301 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      361 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      421 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      481 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      541 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      601 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc
      661 tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac tcactatagg
      721 gagacccaag ctggctagcg tttaaactta agcttgccac catgaaaacg atcatcgccc
      781 tgagctacat cttctgcctg gtattcgcca tgtacaccga tatagagatg aacaggctgg
      841 gaaaggatag cctcatgaac gggaccgagg gcccaaactt ctacgtgcct ttctccaaca
      901 agacgggcgt ggtgcgcagc cccttcgagg ccccgcagta ctacctggcg gagccatggc
      961 agttctccat gctggccgcc tacatgttcc tgctgatcat gcttggcttc cccatcaact
     1021 tcctcacgct gtacgtcttt tatggttata tgaaagaaac aaaacgactc aactacatcc
     1081 tgctcaacct ggccgtggcc gacctcttca tggtgttcgg gggcttcacc accaccctct
     1141 acacctctct gcacgggtac ttcgtctttg ggcccacggg ctgcaacctg gagggcttct
     1201 ttgccacctt gggcggtgaa attgcactgt ggtccttggt ggtcctggcc atcgagcggt
     1261 acgtggtggt gatgtacccc tttcgcttcc atgactgcaa acagaaatat gacgccatca
     1321 tgggcgtcgc cttcacctgg gtcatggctc tggcctgtgc cgcgcccccc ctcgtcggct
     1381 ggtccaggta catcccggag ggcatgcagt gctcgtgcgg gattgactac tacacgcccc
     1441 acgaggaaac caacaatgag tcgttcgtca tctacatgtt cgtggtccac ttcatcatcc
     1501 ccctgattgt catattcttc tgctacggaa aaacggtttt atcactgcaa gataaatatc
     1561 ccatggccca agatcttgga gagaaacaga aagccttgcg catggtgatc atcatggtca
     1621 tcgctttcct aatctgctgg ctgccctacg ctggggtggc gttctacatc ttcacccatc
     1681 agggctctga ctttggcccc atcttcatga ccatcccggc tttctttgcc aagacttctg
     1741 ccgtctacaa ccccgtcatc tacatcatga tgaacaagca gttccgcagg cggctttcaa
     1801 gacaagattt gcatgacagc atccaactcc atgcaaaatc ctttgtgagt aaccatacag
     1861 cttccaccat gacacctgaa ttatgcacag aaaccagcca agtggcgcct gcctaactcg
     1921 agtctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc
     1981 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg
     2041 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc
     2101 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg
     2161 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg
     2221 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca
     2281 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct
     2341 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt
     2401 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac
     2461 gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct
     2521 ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt
     2581 ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac
     2641 aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     2701 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     2761 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2821 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     2881 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     2941 tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     3001 aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag gatgaggatc
     3061 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     3121 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     3181 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     3241 tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     3301 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3361 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     3421 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     3481 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     3541 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat
     3601 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     3661 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     3721 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     3781 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     3841 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     3901 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     3961 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     4021 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     4081 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     4141 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     4201 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     4261 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     4321 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     4381 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     4441 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     4501 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     4561 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     4621 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     4681 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     4741 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     4801 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     4861 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     4921 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     4981 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     5041 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     5101 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     5161 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     5221 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     5281 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     5341 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     5401 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     5461 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     5521 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     5581 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     5641 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     5701 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     5761 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     5821 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     5881 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     5941 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     6001 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     6061 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     6121 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     6181 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     6241 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     6301 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     6361 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
     6421 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
     6481 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgac
//
