LOCUS       Exported                6640 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Expression of a chimeric putative G-protein coupled receptor 
            (Rhodopsin-FFR3; Opto-FFR3; F7; 718).
ACCESSION   .
VERSION     .
KEYWORDS    Opto-FFR3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6640)
  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 6640)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6640
                     /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(2070..2087)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2076..2300
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2346..2774
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2433..2452)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2643..2664
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2783..2803)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2788..3117
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2968..3103
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3030..3049
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3184..3978
                     /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(3238..3257)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3848..3867
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4152..4273
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4189..4208)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4243..4262
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4322..4338)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4322..4338)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4335..4357)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4346..4362
                     /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(4370..4400)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4415..4436
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4553..4570)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4724..5312)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4804..4823)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5483..6343)
                     /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     6106..6125
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6344..6448)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6523..6542)
                     /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 acgtctttgt gggcaagctg cagcgccgcc cggtggccct
     1081 caactacatc ctgctcaacc tggccgtggc cgacctcttc atggtgttcg ggggcttcac
     1141 caccaccctc tacacctctc tgcacgggta cttcgtcttt gggcccacgg gctgcaacct
     1201 ggagggcttc tttgccacct tgggcggtga aattgcactg tggtccttgg tggtcctggc
     1261 catcgagcgg tacgtggtgg tggcccaccc actgtggtac aagacccggc cgaggctggg
     1321 gcaggccatc atgggcgtcg ccttcacctg ggtcatggct ctggcctgtg ccgcgccccc
     1381 cctcgtcggc tggtccaggt acatcccgga gggcatgcag tgctcgtgcg ggattgacta
     1441 ctacacgccc cacgaggaaa ccaacaatga gtcgttcgtc atctacatgt tcgtggtcca
     1501 cttcatcatc cccctgattg tcatattctt ctgctacgga cgcctggtgt ggatcctcgg
     1561 cagagggggc agccaccgcc ggcagaggag ggtggcgcgc atggtgatca tcatggtcat
     1621 cgctttccta atctgctggc tgccctacgc tggggtggcg ttctacatct tcacccatca
     1681 gggctctgac tttggcccca tcttcatgac catcccggct ttctttgcca agacttctgc
     1741 cgtctacaac cccgtcatct acatcatgat gaacaagcag ttccgcgccg actttcatga
     1801 gctgctgagg aggttgtgtg ggctctgggg ccagtggcag caggagagca gcatggagct
     1861 gaaggagcag aagggagggg aggagcagag agcggaccga ccagctgaaa gaaagaccag
     1921 tgaacactca cagggctgtg gaactggtgg ccaggtggcc tgtgctgaaa gcacagaaac
     1981 cagccaagtg gcgcctgcct aagaattcca ccacactgga ctagtggatc cgagctcggt
     2041 accaagctta agtttaaacc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     2101 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     2161 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     2221 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gagaatagca ggcatgctgg
     2281 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
     2341 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2401 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     2461 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     2521 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     2581 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     2641 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     2701 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     2761 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     2821 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     2881 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2941 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     3001 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     3061 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     3121 ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt
     3181 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     3241 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     3301 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     3361 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     3421 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     3481 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     3541 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     3601 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     3661 gaagaacatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     3721 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     3781 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     3841 gacatagcgt tggctacccg tgatattgct gaagaacttg gcggcgaatg ggctgaccgc
     3901 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     3961 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     4021 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     4081 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     4141 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     4201 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4261 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     4321 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4381 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4441 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4501 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttgcgct tcctcgctca
     4561 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     4621 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4681 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4741 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4801 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4861 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4921 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4981 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     5041 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5101 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5161 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5221 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5281 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5341 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5401 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5461 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5521 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5581 gggagggctt accatctggc cccagtgctg caatgatacc gcgggaccca cgctcaccgg
     5641 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5701 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5761 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     5821 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     5881 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     5941 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     6001 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6061 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6121 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6181 ggatcttacc actgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6241 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6301 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6361 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6421 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     6481 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
     6541 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
     6601 gagcaaaatt taagctacaa caaggcaagg cttgaccgac
//
