LOCUS Exported 6625 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Expression of a chimeric G-protein coupled receptor (Rhodopsin-GPR21; Opto-GPR21; A11). ACCESSION . VERSION . KEYWORDS Opto-GPR21 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6625) 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 6625) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6625 /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(2055..2072) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 2061..2285 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 2331..2759 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2418..2437) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2628..2649 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(2768..2788) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 2773..3102 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 2953..3088 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3015..3034 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3169..3963 /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(3223..3242) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3833..3852 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 4137..4258 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4174..4193) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 4228..4247 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(4307..4323) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4307..4323) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4320..4342) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 4331..4347 /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(4355..4385) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4400..4421 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(4538..4555) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4709..5297) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4789..4808) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5468..6328) /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 6091..6110 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6329..6433) /gene="bla" /label=AmpR promoter primer_bind complement(6508..6527) /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 acgtctttca ctgtgcacct ttgttgaacc atcacactct 1081 caactacatc ctgctcaacc tggccgtggc cgacctcttc atggtgttcg ggggcttcac 1141 caccaccctc tacacctctc tgcacgggta cttcgtcttt gggcccacgg gctgcaacct 1201 ggagggcttc tttgccacct tgggcggtga aattgcactg tggtccttgg tggtcctggc 1261 catcgagcgg tacgtggtgg tgactaaacc tttaacctat aatactctgg ttacaccctg 1321 gagagccatc atgggcgtcg ccttcacctg ggtcatggct ctggcctgtg ccgcgccccc 1381 cctcgtcggc tggtccaggt acatcccgga gggcatgcag tgctcgtgcg ggattgacta 1441 ctacacgccc cacgaggaaa ccaacaatga gtcgttcgtc atctacatgt tcgtggtcca 1501 cttcatcatc cccctgattg tcatattctt ctgctacgga aacatcttcc gcatctgcca 1561 acagcacaca aaggacatca gcgaaaggca agcccgcttc agcagccaga gtggggagac 1621 tggggaagtg caggcctgtc ctgataagcg ctatgcccgc atggtgatca tcatggtcat 1681 cgctttccta atctgctggc tgccctacgc tggggtggcg ttctacatct tcacccatca 1741 gggctctgac tttggcccca tcttcatgac catcccggct ttctttgcca agacttctgc 1801 cgtctacaac cccgtcatct acatcatgat gaacaagcag ttccgcagag gactaaagcg 1861 cctctcaggg gctatgtgta cttcttgtgc aagtcagact acagccaacg acccttacac 1921 agttagaagc aaaggccctc ttaatggatg tcatatcaca gaaaccagcc aagtggcgcc 1981 tgcctaagaa ttccaccaca ctggactagt ggatccgagc tcggtaccaa gcttaagttt 2041 aaaccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc 2101 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga 2161 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca 2221 ggacagcaag ggggaggatt gggaagagaa tagcaggcat gctggggatg cggtgggctc 2281 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg 2341 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc 2401 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg 2461 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg 2521 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg 2581 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt 2641 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt 2701 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta 2761 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga 2821 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc 2881 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc 2941 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc 3001 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag 3061 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt 3121 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa 3181 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg 3241 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc 3301 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca 3361 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc 3421 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca 3481 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat 3541 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca 3601 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga acatcagggg 3661 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc 3721 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct 3781 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct 3841 acccgtgata ttgctgaaga acttggcggc gaatgggctg accgcttcct cgtgctttac 3901 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc 3961 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag 4021 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg 4081 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact 4141 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata 4201 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc 4261 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc 4321 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt 4381 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc 4441 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg 4501 ggagaggcgg tttgcgtatt gggcgctctt gcgcttcctc gctcactgac tcgctgcgct 4561 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca 4621 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga 4681 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc 4741 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg 4801 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat 4861 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt 4921 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc 4981 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg 5041 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg 5101 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg 5161 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg 5221 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca 5281 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga 5341 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga 5401 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt 5461 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt 5521 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat 5581 ctggccccag tgctgcaatg ataccgcggg acccacgctc accggctcca gatttatcag 5641 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct 5701 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt 5761 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg 5821 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca 5881 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt 5941 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat 6001 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac 6061 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa 6121 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccactgt 6181 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt 6241 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa 6301 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt 6361 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa 6421 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc 6481 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca 6541 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc 6601 tacaacaagg caaggcttga ccgac //