LOCUS       sequence-209707-        6841 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of a chimeric putative G-protein coupled receptor 
            (Rhodopsin-A2AR; Opto-A2AR; F8; 719).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6841)
  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 6841)
  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-A2AR
FEATURES             Location/Qualifiers
     source          1..6841
                     /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             821..853
                     /codon_start=1
                     /product="epitope tag from vesicular stomatitis virus G 
                     protein"
                     /label=VSV-G tag
                     /translation="YTDIEMNRLGK"
     CDS             878..916
                     /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(2271..2288)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2277..2501
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2547..2975
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2634..2653)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2844..2865
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2984..3004)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2989..3318
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3169..3304
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3231..3250
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3385..4179
                     /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(3439..3458)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4049..4068
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4353..4474
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4390..4409)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4444..4463
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4523..4539)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4523..4539)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4536..4558)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4547..4563
                     /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(4571..4577,4578..4595,4596..4601))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4616..4637
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4754..4771)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4925..5513)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5005..5024)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5684..6475,6476..6544))
                     /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     6307..6326
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6545..6649)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6724..6743)
                     /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 gagacacaag ctggctagcg tttaaacggg ccctctagac tcgagcacca tgaaaacgat
      781 catcgccctg agctacatct tctgcctggt attcgccatg tacaccgata tagagatgaa
      841 caggctggga aaggatagcc tcatgaacgg gaccgagggc ccaaacttct acgtgccttt
      901 ctccaacaag acgggcgtgg tgcgcagccc cttcgaggcc ccgcagtact acctggcgga
      961 gccatggcag ttctccatgc tggccgccta catgttcctg ctgatcatgc ttggcttccc
     1021 catcaacttc ctcacgctgt acgtcgtgtg gctcaacagc aacctgcaga acgtcctcaa
     1081 ctacatcctg ctcaacctgg ccgtggccga cctcttcatg gtgttcgggg gcttcaccac
     1141 caccctctac acctctctgc acgggtactt cgtctttggg cccacgggct gcaacctgga
     1201 gggcttcttt gccaccttgg gcggtgaaat tgcactgtgg tccttggtgg tcctggccat
     1261 cgagcggtac gtggtggtgc gcatcccgct ccggtacaat ggcttggtga ccggcacgag
     1321 ggccatcatg ggcgtcgcct tcacctgggt catggctctg gcctgtgccg cgccccccct
     1381 cgtcggctgg tccaggtaca tcccggaggg catgcagtgc tcgtgcggga ttgactacta
     1441 cacgccccac gaggaaacca acaatgagtc gttcgtcatc tacatgttcg tggtccactt
     1501 catcatcccc ctgattgtca tattcttctg ctacggacgg atcttcctgg cggcgcgacg
     1561 acagctgaag cagatggaga gccagcctct gccgggggag cgggcacggt ccacactgca
     1621 gaaggaggtc catgctcgca tggtgatcat catggtcatc gctttcctaa tctgctggct
     1681 gccctacgct ggggtggcgt tctacatctt cacccatcag ggctctgact ttggccccat
     1741 cttcatgacc atcccggctt tctttgccaa gacttctgcc gtctacaacc ccgtcatcta
     1801 catcatgatg aacaagcagt tccgccagac cttccgcaag atcattcgca gccacgtcct
     1861 gaggcagcaa gaacctttca aggcagctgg caccagtgcc cgggtcttgg cagctcatgg
     1921 cagtgacgga gagcaggtca gcctccggct caacggccac ccgccaggag tgtgggccaa
     1981 cggcagtgct ccccaccctg agcggaggcc caatggctat gccctggggc tggtgagtgg
     2041 agggagtgcc caagagtccc aggggaacac gggcctccca gacgtggagc tccttagcca
     2101 tgagctcaag ggagtgtgcc cagagccccc tggcctagat gaccccctgg cccaggatgg
     2161 agcaggagtg tccacagaaa ccagccaagt ggcgcctgcc taagaattcc accacactgg
     2221 actagtggat ccgagctcgg taccaagctt aagtttaaac cgctgatcag cctcgactgt
     2281 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     2341 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     2401 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2461 agagaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     2521 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     2581 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2641 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2701 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2761 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2821 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2881 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     2941 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     3001 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3061 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3121 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3181 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3241 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3301 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     3361 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     3421 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3481 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3541 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3601 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3661 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3721 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3781 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3841 gtcgatcagg atgatctgga cgaagaacat caggggctcg cgccagccga actgttcgcc
     3901 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     3961 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4021 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagaactt
     4081 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     4141 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     4201 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     4261 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     4321 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     4381 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4441 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4501 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4561 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4621 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4681 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4741 gctcttgcgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4801 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4861 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4921 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4981 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5041 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5101 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5161 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5221 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5281 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5341 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     5401 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5461 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5521 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5581 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5641 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5701 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5761 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5821 cgcgggaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5881 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5941 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     6001 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     6061 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     6121 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6181 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6241 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6301 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6361 cttcggggcg aaaactctca aggatcttac cactgttgag atccagttcg atgtaaccca
     6421 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6481 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6541 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6601 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6661 gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct
     6721 cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt
     6781 ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga
     6841 c
//
