LOCUS Exported 6103 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses NanoLuc-beta-2AR-AsLOV2(V416L) in mammalian cells. ACCESSION . VERSION . KEYWORDS pAAV-gLuc sp-NanoLuc-HA-beta-2AR-NNES-AsLOV2(V416L) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6103) TITLE Luciferase-LOV BRET enables versatile and specific transcriptional readout of cellular protein-protein interactions JOURNAL Unpublished REFERENCE 2 (bases 1 to 6103) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6103 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 6..135 /label=AAV2 ITR (alternate) /note="Functional equivalent of wild-type AAV2 ITR" enhancer 206..509 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 510..713 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 664..684 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 758..777 /label=T7 /note="T7 promoter, forward primer" promoter 758..776 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" regulatory 789..798 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 846..1355 /codon_start=1 /product="NanoLuc(R) luciferase" /label=Nluc /note="human codon-optimized" /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR LCERILA" CDS 1356..1382 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" primer_bind complement(3129..3146) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3135..3342 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" repeat_region 3371..3511 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" repeat_region 3371..3500 /label=AAV2 ITR rep_origin 3586..4041 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3673..3692) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3883..3904 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(4058..4077) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4177..4199 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4237..4255) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 4323..4427 /gene="bla" /label=AmpR promoter CDS 4428..5288 /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 complement(4646..4665) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5459..6047 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5948..5967 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 catgtcctgc aggcagctgc gcgctcgctc gctcactgag gccgcccggg cgtcgggcga 61 cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg gccaactcca 121 tcactagggg ttcctgcggc cgcacgcgtt gtacatagta atcaattacg gggtcattag 181 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct 241 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc 301 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg 361 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat 421 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca 481 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc 541 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga 601 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat 661 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc 721 taactagaga acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga 781 cccagcttgc caccatggga gtcaaagttc tgtttgccct gatctgcatc gctgtggccg 841 aggccgtctt cacactcgaa gatttcgttg gggactggcg acagacagcc ggctacaacc 901 tggaccaagt ccttgaacag ggaggtgtgt ccagtttgtt tcagaatctc ggggtgtccg 961 taactccgat ccaaaggatt gtcctgagcg gtgaaaatgg gctgaagatc gacatccatg 1021 tcatcatccc gtatgaaggt ctgagcggcg accaaatggg ccagatcgaa aaaattttta 1081 aggtggtgta ccctgtggat gatcatcact ttaaggtgat cctgcactat ggcacactgg 1141 taatcgacgg ggttacgccg aacatgatcg actatttcgg acggccgtat gaaggcatcg 1201 ccgtgttcga cggcaaaaag atcactgtaa cagggaccct gtggaacggc aacaaaatta 1261 tcgacgagcg cctgatcaac cccgacggct ccctgctgtt ccgagtaacc atcaacggag 1321 tgaccggctg gcggctgtgc gaacgcattc tggcgtatcc gtacgacgta ccagactacg 1381 cagggcaacc cgggaacggc agcgccttct tgctggcacc caatagaagc catgcgccgg 1441 accacgacgt cacgcagcag agggacgagg tgtgggtggt gggcatgggc atcgtcatgt 1501 ctctcatcgt cctggccatc gtgtttggca atgtgctggt catcacagcc attgccaagt 1561 tcgagcgtct gcagacggtc accaactact tcatcacttc actggcctgt gctgatctgg 1621 tcatgggcct ggcagtggtg ccctttgggg ccgcccatat tcttatgaaa atgtggactt 1681 ttggcaactt ctggtgcgag ttttggactt ccattgatgt gctgtgcgtc acggccagca 1741 ttgagaccct gtgcgtgatc gcagtggatc gctactttgc cattacttca cctttcaagt 1801 accagagcct gctgaccaag aataaggccc gggtgatcat tctgatggtg tggattgtgt 1861 caggccttac ctccttcttg cccattcaga tgcactggta ccgggccacc caccaggaag 1921 ccatcaactg ctatgccaat gagacctgct gtgacttctt cacgaaccaa gcctatgcca 1981 ttgcctcttc catcgtgtcc ttctacgttc ccctggtgat catggtcttc gtctactcca 2041 gggtctttca ggaggccaaa aggcagctcc agaagattga caaatctgag ggccgcttcc 2101 atgtccagaa ccttagccag gtggagcagg atgggcggac ggggcatgga ctccgcagat 2161 cttccaagtt ctgcttgaag gagcacaaag ccctcaagac gttaggcatc atcatgggca 2221 ctttcaccct ctgctggctg cccttcttca tcgttaacat tgtgcatgtg atccaggata 2281 acctcatccg taaggaagtt tacatcctcc taaattggat aggctatgtc aattctggtt 2341 tcaatcccct tatctactgc cggagcccag atttcaggat tgccttccag gagcttctgt 2401 gcctgcgcag gtcttctttg aaggcctatg ggaatggcta ctccagcaac ggcaacacag 2461 gggagcagag tggatatcac gtggaacagg agaaagaaaa taaactgctg tgtgaagacc 2521 tcccaggcac ggaagacttt gtgggccatc aaggtactgt gcctagcgat aacattgatt 2581 cacaagggag gaattgtagt acaaatgact cactgctgga gctggccgag aagctggccg 2641 gcctggacat caacggtgga gctagcttgg ctactacact tgaacgtatt gagaagaact 2701 ttctcattac tgacccaaga ttgccagata atcccattat attcgcgtcc gatagtttct 2761 tgcagttgac agaatatagc cgtgaagaaa ttttgggaag aaactgcagg tttctacaag 2821 gtcctgaaac tgatcgcgcg acagtgagaa aaattagaga tgccatagat aaccaaacag 2881 aggtcactgt tcagctgatt aattatacaa agagtggtaa aaagttctgg aacctctttc 2941 acttgcagcc tatgcgagat cagaagggag atgtccagta ctttattggg gttcagttgg 3001 atggaactga gcatgtccga gatgctgccg agagagaggg agtcatgctg attaagaaaa 3061 ctgcagaaaa tattgatgag gcggcaaaag aactttaaga attcctcgag ctagagctcg 3121 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt 3181 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat 3241 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag 3301 caagggggag gattgggaag acaatagcag gcatgctggg gaggtaacca cgtgcggacc 3361 gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg 3421 ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca 3481 gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt tctccttacg 3541 catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg 3601 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg 3661 ccttagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc 3721 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc 3781 tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga 3841 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa 3901 ctggaacaac actcaactct atctcgggct attcttttga tttataaggg attttgccga 3961 tttcggtcta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca 4021 aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat 4081 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc 4141 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt 4201 tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat 4261 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg 4321 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga 4381 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac 4441 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc 4501 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca 4561 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc 4621 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg 4681 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac 4741 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca 4801 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg 4861 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac 4921 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg 4981 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat 5041 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg 5101 ctggctggtt tattgctgat aaatctggag ccggtgagcg tggaagccgc ggtatcattg 5161 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc 5221 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc 5281 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt 5341 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt 5401 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt 5461 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag 5521 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca 5581 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca 5641 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg 5701 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg 5761 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct 5821 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga 5881 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc 5941 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg 6001 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg 6061 cggccttttt acggttcctg gccttttgct ggccttttgc tca //