LOCUS       Exported                6712 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expression of a chimeric G-protein coupled receptor 
            (Rhodopsin-GPR27; Opto-GPR27; B3).
ACCESSION   .
VERSION     .
KEYWORDS    Opto-GPR27
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6712)
  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 6712)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6712
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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(2142..2159)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2148..2372
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2418..2846
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2505..2524)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2715..2736
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2855..2875)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2860..3189
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3040..3175
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3102..3121
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3256..4050
                     /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(3310..3329)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3920..3939
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4224..4345
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4261..4280)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4315..4334
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4394..4410)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4394..4410)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4407..4429)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4418..4434
                     /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(4442..4472)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4487..4508
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4625..4642)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4796..5384)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4876..4895)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5555..6415)
                     /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     6178..6197
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6416..6520)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6595..6614)
                     /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 acgtcatcgt gcgggagcgc agcctgcacc gcgccctcaa
     1081 ctacatcctg ctcaacctgg ccgtggccga cctcttcatg gtgttcgggg gcttcaccac
     1141 caccctctac acctctctgc acgggtactt cgtctttggg cccacgggct gcaacctgga
     1201 gggcttcttt gccaccttgg gcggtgaaat tgcactgtgg tccttggtgg tcctggccat
     1261 cgagcggtac gtggtggtgg cgcaccaccg cttctatgca gagcgcctgg ccggctggcc
     1321 gtgcgccgcc atcatgggcg tcgccttcac ctgggtcatg gctctggcct gtgccgcgcc
     1381 ccccctcgtc ggctggtcca ggtacatccc ggagggcatg cagtgctcgt gcgggattga
     1441 ctactacacg ccccacgagg aaaccaacaa tgagtcgttc gtcatctaca tgttcgtggt
     1501 ccacttcatc atccccctga ttgtcatatt cttctgctac ggacacgacc gccgcaagat
     1561 gcggcccgcg cgcctggtgc ccgccgtcag ccacgactgg accttccacg gcccgggcgc
     1621 caccggccag gccgccgcca actggacggc gggcttcggc cgcgggccca cgccgcccgc
     1681 gcttgtgggc atccggcccg cagggccggg ccgcggcgcg cgccgcctcc tcgtgctgga
     1741 agaatttaag acggagaaga ggctgtgccg catggtgatc atcatggtca tcgctttcct
     1801 aatctgctgg ctgccctacg ctggggtggc gttctacatc ttcacccatc agggctctga
     1861 ctttggcccc atcttcatga ccatcccggc tttctttgcc aagacttctg ccgtctacaa
     1921 ccccgtcatc tacatcatga tgaacaagca gttccgcgac tgcttcaggg cccagttccc
     1981 ctgctgccag agcccccgga ccacccaggc gacccatccc tgcgacctga aaggcattgg
     2041 tttaacagaa accagccaag tggcgcctgc ctaagaattc caccacactg gactagtgga
     2101 tccgagctcg gtaccaagct taagtttaaa ccgctgatca gcctcgactg tgccttctag
     2161 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     2221 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     2281 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagagaatag
     2341 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     2401 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     2461 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     2521 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     2581 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     2641 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     2701 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     2761 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     2821 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     2881 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2941 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     3001 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     3061 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     3121 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     3181 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     3241 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     3301 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     3361 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     3421 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     3481 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     3541 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     3601 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     3661 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     3721 gatgatctgg acgaagaaca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     3781 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     3841 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     3901 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagaact tggcggcgaa
     3961 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     4021 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     4081 aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt
     4141 tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca
     4201 tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa
     4261 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     4321 tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct
     4381 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     4441 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     4501 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     4561 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttgcg
     4621 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4681 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4741 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4801 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4861 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4921 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4981 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5041 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5101 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5161 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5221 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5281 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5341 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5401 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5461 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     5521 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     5581 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     5641 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgggacc
     5701 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     5761 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     5821 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     5881 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     5941 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     6001 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     6061 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     6121 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     6181 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     6241 gaaaactctc aaggatctta ccactgttga gatccagttc gatgtaaccc actcgtgcac
     6301 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     6361 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     6421 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     6481 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     6541 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat
     6601 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc
     6661 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg ac
//
