LOCUS Exported 7625 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expression of G protein-coupled receptors for PRESTO-Tango: parallel receptorome expression and screening via transcriptional output, with transcriptional activation following arrestin translocation. ACCESSION . VERSION . KEYWORDS GPR3-Tango SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7625) AUTHORS Kroeze WK, Sassano MF, Huang XP, Lansu K, McCorvy JD, Giguere PM, Sciaky N, Roth BL TITLE PRESTO-Tango as an open-source resource for interrogation of the druggable human GPCRome. JOURNAL Nat Struct Mol Biol. 2015 May;22(5):362-9. doi: 10.1038/nsmb.3014. Epub 2015 Apr 20. PUBMED 25895059 REFERENCE 2 (bases 1 to 7625) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7625 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(64..84) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 69..398 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 249..384 /label=SV40 ori /note="SV40 origin of replication" primer_bind 311..330 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 465..1259 /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(519..538) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 1129..1148 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 1433..1554 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1470..1489) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1524..1543 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(1603..1619) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1603..1619) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1616..1638) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1627..1643 /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(1651..1681) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1696..1717 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(1834..1851) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(2005..2593) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2085..2104) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(2764..3624) /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 3387..3406 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(3625..3729) /gene="bla" /label=AmpR promoter primer_bind complement(3804..3823) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 3995..4374 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4375..4578 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4529..4549 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 4623..4642 /label=T7 /note="T7 promoter, forward primer" promoter 4623..4641 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 5943..6944 /codon_start=1 /product="tetracycline-controlled transactivator, comprising a fusion of the tetracycline repressor TetR with the C-terminal activation domain of herpes simplex virus VP16" /label=tTA /note="In the Tet-Off(R) system, tTA binds to the Tet-responsive element and stimulates transcription only in the absence of tetracycline or doxycycline." /translation="RLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVKN KRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRPT EKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTDS MPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTIEG LLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHADA LDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG" primer_bind complement(6010..6029) /label=Tet-R /note="Tetracycline resistance gene, reverse primer" primer_bind complement(6976..6993) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 6982..7206 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 7252..55 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(7339..7358) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 7549..7570 /label=F1ori-F /note="F1 origin, forward primer" ORIGIN 1 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc 61 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta 121 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag 181 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa 241 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac 301 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt 361 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat 421 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg 481 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac 541 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg 601 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc 661 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg 721 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc 781 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc 841 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta 901 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg 961 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg 1021 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat 1081 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc 1141 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta 1201 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag 1261 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt 1321 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg 1381 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt 1441 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc 1501 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt 1561 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt 1621 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa 1681 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc 1741 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag 1801 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt 1861 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga 1921 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg 1981 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa 2041 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt 2101 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct 2161 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct 2221 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc 2281 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt 2341 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc 2401 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat 2461 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa 2521 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa 2581 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga 2641 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct 2701 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga 2761 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc 2821 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg 2881 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat 2941 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat 3001 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg 3061 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc 3121 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa 3181 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc 3241 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt 3301 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag 3361 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt 3421 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag 3481 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac 3541 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc 3601 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca 3661 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg 3721 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc 3781 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat 3841 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca 3901 acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg 3961 ctgcttcgcg atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa 4021 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa 4081 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata 4141 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag 4201 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc 4261 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta 4321 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg 4381 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt 4441 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca 4501 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag 4561 gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa 4621 attaatacga ctcactatag ggagacccaa gctggctagc gtttaaactt aagcttggta 4681 ccgagctcgg atccactagt ccagtgtggt ggaattctgc agatatccag cacagtggcg 4741 gccgcgccac catgaagacg atcatcgccc tgagctacat cttctgcctg gtattcgccg 4801 actacaagga cgatgatgac gccagcatcg atatgatgtg gggggctggg tctccactgg 4861 cctggctgtc tgctggtagc ggaaacgtga acgttagttc cgtgggaccc gctgaggggc 4921 caacgggccc agcagcgcct ctccctagtc ccaaagcttg ggacgtcgtg ttgtgcattt 4981 caggcaccct ggtttcctgt gagaacgccc ttgtggtggc cattatcgtt ggcacaccag 5041 cgtttagggc ccctatgttc ctgcttgtgg gctccctggc cgttgcggat ttgctggctg 5101 ggctcggtct cgtgttgcac tttgctgcag tattttgtat cggaagcgcc gagatgtccc 5161 tcgtgctcgt gggcgtgctt gctatggctt tcacagcttc aatcggcagc ttgctcgcca 5221 ttactgtaga ccgctacctg tccctgtaca acgcattgac ctactactca gaaaccacag 5281 tcacaagaac ctacgtaatg ctggcactgg tttggggtgg cgcactgggc cttggacttc 5341 ttcccgtgtt ggcctggaac tgtcttgatg ggctcactac ctgtggagtt gtctaccctt 5401 tgagcaagaa tcatctggta gtgctcgcca tagccttttt tatggtgttc gggattatgc 5461 ttcagctgta tgcccaaatt tgcaggatcg tttgtaggca cgcacagcag attgctttgc 5521 agaggcacct gcttccagct agccactatg tggcgactcg gaaaggcatc gcaaccttgg 5581 ccgtcgtgtt gggggctttt gccgcatgct ggctgccctt tacggtgtat tgcctgctgg 5641 gagatgccca tagtccccct ctttacactt atctgacgct cctgcctgcg acgtataact 5701 ccatgattaa tcctatcatc tacgccttcc ggaatcagga tgtacagaag gtgctctggg 5761 cagtctgttg ctgctgtagc tcttcaaaaa tccccttcag gtcccggagc ccctccgatg 5821 tcatcgatac cggtggacgc accccaccca gcctgggtcc ccaagatgag tcctgcacca 5881 ccgccagctc ctccctggcc aaggacactt catcgaccgg tgagaacctg tacttccagc 5941 taagattaga taaaagtaaa gtgattaaca gcgcattaga gctgcttaat gaggtcggaa 6001 tcgaaggttt aacaacccgt aaactcgccc agaagctagg tgtagagcag cctacattgt 6061 attggcatgt aaaaaataag cgggctttgc tcgacgcctt agccattgag atgttagata 6121 ggcaccatac tcacttttgc cctttagaag gggaaagctg gcaagatttt ttacgtaata 6181 acgctaaaag ttttagatgt gctttactaa gtcatcgcga tggagcaaaa gtacatttag 6241 gtacacggcc tacagaaaaa cagtatgaaa ctctcgaaaa tcaattagcc tttttatgcc 6301 aacaaggttt ttcactagag aatgcattat atgcactcag cgctgtgggg cattttactt 6361 taggttgcgt attggaagat caagagcatc aagtcgctaa agaagaaagg gaaacaccta 6421 ctactgatag tatgccgcca ttattacgac aagctatcga attatttgat caccaaggtg 6481 cagagccagc cttcttattc ggccttgaat tgatcatatg cggattagaa aaacaactta 6541 aatgtgaaag tgggtccgcg tacagccgcg cgcgtacgaa aaacaattac gggtctacca 6601 tcgagggcct gctcgatctc ccggacgacg acgcccccga agaggcgggg ctggcggctc 6661 cgcgcctgtc ctttctcccc gcgggacaca cgcgcagact gtcgacggcc cccccgaccg 6721 atgtcagcct gggggacgag ctccacttag acggcgagga cgtggcgatg gcgcatgccg 6781 acgcgctaga cgatttcgat ctggacatgt tgggggacgg ggattccccg ggtccgggat 6841 ttacccccca cgactccgcc ccctacggcg ctctggatat ggccgacttc gagtttgagc 6901 agatgtttac cgatgccctt ggaattgacg agtacggtgg gtagtctaga gggcccgttt 6961 aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct 7021 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg 7081 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc 7141 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct 7201 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct 7261 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg 7321 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg 7381 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac 7441 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct 7501 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt 7561 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt 7621 tgccg //