LOCUS Exported 7183 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expression of a chimeric putative G-protein coupled receptor (Rhodopsin-M3R; Opto-M3R; H1; 724). ACCESSION . VERSION . KEYWORDS Opto-M3R SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7183) 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 7183) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7183 /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 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(2613..2630) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 2619..2843 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 2889..3317 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2976..2995) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3186..3207 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(3326..3346) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3331..3660 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3511..3646 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3573..3592 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3727..4521 /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(3781..3800) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4391..4410 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 4695..4816 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4732..4751) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4786..4805 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(4865..4881) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4865..4881) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4878..4900) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4889..4905 /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(4913..4943) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4958..4979 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5096..5113) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5267..5855) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5347..5366) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6026..6886) /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 6649..6668 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6887..6991) /gene="bla" /label=AmpR promoter primer_bind complement(7066..7085) /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 aaggtcaaca agcagctgaa gacggtcctc aactacatcc 1081 tgctcaacct ggccgtggcc gacctcttca tggtgttcgg gggcttcacc accaccctct 1141 acacctctct gcacgggtac ttcgtctttg ggcccacggg ctgcaacctg gagggcttct 1201 ttgccacctt gggcggtgaa attgcactgt ggtccttggt ggtcctggcc atcgagcggt 1261 acgtggtggt gacgaggccg ctcacgtacc gagccaaacg aacaacaaag agagccatca 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 ggatctataa ggaaactgaa aagcgtacca 1561 aagagcttgc tggcctgcaa gcctctggga cagaggcaga gacagaaaac tttgtccacc 1621 ccacgggcag ttctcgaagc tgcagcagtt acgaacttca acagcaaagc atgaaacgct 1681 ccaacaggag gaagtatggc cgctgccact tctggttcac aaccaagagc tggaaaccca 1741 gctccgagca gatggaccaa gaccacagca gcagtgacag ttggaacaac aatgatgctg 1801 ctgcctccct ggagaactcc gcctcctccg acgaggagga cattggctcc gagacgagag 1861 ccatctactc catcgtgctc aagcttccgg gtcacagcac catcctcaac tccaccaagt 1921 taccctcatc ggacaacctg caggtgcctg aggaggagct ggggatggtg gacttggaga 1981 ggaaagccga caagctgcag gcccagaaga gcgtggacga tggaggcagt tttccaaaaa 2041 gcttctccaa gcttcccatc cagctagagt cagccgtgga cacagctaag acttctgacg 2101 tcaactcctc agtgggtaag agcacggcca ctctacctct gtccttcaag gaagccactc 2161 tggccaagag gtttgctctg aagaccagaa gtcagatcac taagcggaaa aggatgtccc 2221 tggtcaagga gaagaaagcg gcccgcatgg tgatcatcat ggtcatcgct ttcctaatct 2281 gctggctgcc ctacgctggg gtggcgttct acatcttcac ccatcagggc tctgactttg 2341 gccccatctt catgaccatc ccggctttct ttgccaagac ttctgccgtc tacaaccccg 2401 tcatctacat catgatgaac aagcagttcc gcaccacttt caagatgctg ctgctgtgcc 2461 agtgtgacaa aaaaaagagg cgcaagcagc agtaccagca gagacagtcg gtcatttttc 2521 acaagcgcgc acccgagcag gccttgacag aaaccagcca agtggcgcct gcctaactcg 2581 agtctagagg gcccgtttaa accgctgatc agcctcgact gtgccttcta gttgccagcc 2641 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt 2701 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct 2761 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc 2821 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg 2881 gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag 2941 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt 3001 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt 3061 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg 3121 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt 3181 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt 3241 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca 3301 aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca 3361 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt 3421 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca 3481 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc 3541 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct 3601 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa 3661 aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg 3721 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg 3781 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg 3841 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat 3901 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca 3961 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg 4021 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat 4081 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa 4141 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg 4201 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg 4261 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg 4321 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat 4381 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac 4441 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc 4501 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc 4561 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg 4621 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt 4681 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca 4741 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac 4801 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat 4861 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac 4921 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa 4981 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat 5041 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc 5101 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg 5161 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag 5221 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc 5281 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag 5341 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga 5401 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc 5461 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg 5521 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt 5581 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca 5641 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca 5701 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag 5761 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca 5821 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg 5881 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa 5941 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta 6001 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag 6061 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga 6121 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac 6181 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc 6241 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta 6301 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac 6361 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat 6421 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa 6481 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg 6541 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag 6601 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc 6661 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct 6721 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat 6781 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg 6841 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc 6901 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta 6961 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg 7021 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga 7081 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg 7141 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gac //