LOCUS Exported 7691 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 CXCR4-Tango SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7691) 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 7691) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7691 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..380 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 381..584 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 535..555 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 629..648 /label=T7 /note="T7 promoter, forward primer" promoter 629..647 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" CDS 2015..3016 /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(2082..2101) /label=Tet-R /note="Tetracycline resistance gene, reverse primer" primer_bind complement(3048..3065) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3054..3278 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 3324..3752 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3411..3430) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3621..3642 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(3761..3781) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3766..4095 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3946..4081 /label=SV40 ori /note="SV40 origin of replication" primer_bind 4008..4027 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 4162..4956 /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(4216..4235) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4826..4845 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 5130..5251 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5167..5186) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5221..5240 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(5300..5316) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5300..5316) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5313..5335) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5324..5340 /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(5348..5378) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5393..5414 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5531..5548) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5702..6290) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5782..5801) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6461..7321) /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 7084..7103 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7322..7426) /gene="bla" /label=AmpR promoter primer_bind complement(7501..7520) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc 61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca 121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga 181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc 241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct 301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat 361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc 421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt 481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa 541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag 601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg 661 gctagcgttt aaacttaagc ttggtaccga gctcggatcc actagtccag tgtggtggaa 721 ttctgcagat atccagcaca gtggcggccg cgccaccatg aagacgatca tcgccctgag 781 ctacatcttc tgcctggtat tcgccgacta caaggacgat gatgacgcca gcatcgatat 841 ggaggggata tcaatctaca catcagataa ttacacggaa gagatgggtt ccggcgatta 901 cgactctatg aaagagccgt gttttagaga ggaaaacgcg aactttaaca agatctttct 961 ccccaccatc tacagcatca tctttctcac aggcatcgta gggaacggcc tggtcatcct 1021 ggtgatggga taccaaaaga agctgaggtc aatgaccgac aagtataggc tccatctgtc 1081 cgtggccgac ctcctgtttg tgattaccct gcctttttgg gcagttgacg ctgtcgctaa 1141 ttggtacttc ggcaacttcc tctgtaaggc agtgcacgtt atctacactg tgaatcttta 1201 tagttccgtc ttgatcttgg cctttatcag tctcgacagg tatttggcga ttgtgcacgc 1261 taccaactca caacgaccta gaaaactcct ggctgagaag gtggtttacg ttggtgtgtg 1321 gattccagct ctcctgttga caataccaga cttcattttc gctaatgtga gcgaggccga 1381 tgacagatac atttgtgacc gattttaccc aaacgatctg tgggtagtgg tatttcagtt 1441 ccaacacatt atggtcgggc tgatcttgcc cggcattgtc atactgtctt gctactgcat 1501 cattatttct aagctgtcac actcaaaagg ccaccaaaag aggaaggctc tgaaaacaac 1561 ggtgatcctg atactggcct tcttcgcatg ttggctgccc tactatatcg gcatcagcat 1621 tgactcattt atactcctgg aaattatcaa gcagggctgc gagttcgaga acaccgttca 1681 taagtggatt tctataaccg aggccctcgc cttctttcac tgttgtttga atccgattct 1741 ctacgcgttt cttggcgcca aatttaaaac aagcgcccaa catgcactga catcagtgtc 1801 tagggggagc tctctgaaaa tcctctccaa gggaaaacga ggcggacata gcagtgtcag 1861 cactgagtcc gaatccagct catttcatag ctctatcgat accggtggac gcaccccacc 1921 cagcctgggt ccccaagatg agtcctgcac caccgccagc tcctccctgg ccaaggacac 1981 ttcatcgacc ggtgagaacc tgtacttcca gctaagatta gataaaagta aagtgattaa 2041 cagcgcatta gagctgctta atgaggtcgg aatcgaaggt ttaacaaccc gtaaactcgc 2101 ccagaagcta ggtgtagagc agcctacatt gtattggcat gtaaaaaata agcgggcttt 2161 gctcgacgcc ttagccattg agatgttaga taggcaccat actcactttt gccctttaga 2221 aggggaaagc tggcaagatt ttttacgtaa taacgctaaa agttttagat gtgctttact 2281 aagtcatcgc gatggagcaa aagtacattt aggtacacgg cctacagaaa aacagtatga 2341 aactctcgaa aatcaattag cctttttatg ccaacaaggt ttttcactag agaatgcatt 2401 atatgcactc agcgctgtgg ggcattttac tttaggttgc gtattggaag atcaagagca 2461 tcaagtcgct aaagaagaaa gggaaacacc tactactgat agtatgccgc cattattacg 2521 acaagctatc gaattatttg atcaccaagg tgcagagcca gccttcttat tcggccttga 2581 attgatcata tgcggattag aaaaacaact taaatgtgaa agtgggtccg cgtacagccg 2641 cgcgcgtacg aaaaacaatt acgggtctac catcgagggc ctgctcgatc tcccggacga 2701 cgacgccccc gaagaggcgg ggctggcggc tccgcgcctg tcctttctcc ccgcgggaca 2761 cacgcgcaga ctgtcgacgg cccccccgac cgatgtcagc ctgggggacg agctccactt 2821 agacggcgag gacgtggcga tggcgcatgc cgacgcgcta gacgatttcg atctggacat 2881 gttgggggac ggggattccc cgggtccggg atttaccccc cacgactccg ccccctacgg 2941 cgctctggat atggccgact tcgagtttga gcagatgttt accgatgccc ttggaattga 3001 cgagtacggt gggtagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc 3061 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg 3121 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag 3181 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga 3241 caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag 3301 ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt 3361 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc 3421 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg 3481 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta 3541 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt 3601 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat 3661 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa 3721 tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg 3781 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag 3841 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg 3901 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact 3961 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag 4021 gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc 4081 ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag 4141 acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc 4201 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat 4261 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg 4321 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg 4381 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta 4441 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta 4501 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc 4561 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc 4621 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg 4681 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg 4741 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt 4801 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc 4861 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc 4921 atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga 4981 ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg 5041 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg 5101 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca 5161 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt 5221 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct 5281 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa 5341 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga 5401 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt 5461 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct 5521 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat 5581 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga 5641 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt 5701 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt 5761 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc 5821 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa 5881 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct 5941 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta 6001 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg 6061 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc 6121 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta 6181 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg 6241 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt 6301 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg 6361 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta 6421 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg 6481 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg 6541 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc 6601 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg 6661 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg 6721 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag 6781 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat 6841 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc 6901 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc 6961 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa 7021 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac 7081 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt 7141 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc 7201 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa 7261 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca 7321 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat 7381 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa 7441 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag 7501 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga 7561 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa 7621 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag 7681 atatacgcgt t //