LOCUS Exported 5751 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION mammalian expression and localization. ACCESSION . VERSION . KEYWORDS EGFP-Nsp15 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5751) AUTHORS Miserey-Lenkei S, Trajkovic K, D'Ambrosio JM, Patel AJ, Copic A, Mathur P, Schauer K, Goud b, Albanese V, Gautier R, Subra M, Kovacs D, Barelli H, Antonny B TITLE A comprehensive library of fluorescent constructs of SARS-CoV-2 proteins and their initial characterization in different cell types JOURNAL bioRxiv 423586 REFERENCE 2 (bases 1 to 5751) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5751 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 1..204 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 155..175 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 243..252 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 249..965 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 249..965 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(294..315) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(555..574) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 902..923 /label=EGFP-C /note="EGFP, forward primer" polyA_signal 2175..2296 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2212..2231) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2266..2285 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2303..2758) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2440..2461) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 2652..2671 /label=F1ori-R /note="F1 origin, reverse primer" promoter 2785..2889 /gene="bla" /label=AmpR promoter promoter 2891..3248 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(2914..2934) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3099..3234 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3161..3180 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3283..4077 /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 GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(3337..3356) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3947..3966 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4265..4284) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4309..4356 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 4685..5273 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5174..5193 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 5448..5751 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" ORIGIN 1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt 61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac 121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg 181 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg 241 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg 301 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg 361 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct 421 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc 481 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca 541 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg 601 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc 661 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc 721 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc 781 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 841 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 901 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 961 acaagtccgg actcagatct cgagctcaag cttcgaattc tatgagtctt gaaaacgtcg 1021 ccttcaacgt agtaaataag ggccactttg atggtcaaca gggagaagta cccgtgtcta 1081 tcatcaataa caccgtttac acgaaggttg acggcgttga tgttgaactg ttcgaaaata 1141 agacgacact cccggtgaac gtagctttcg agctttgggc gaagcgaaat ataaaacctg 1201 tccctgaagt taaaattctc aataatcttg gggtagacat cgcggctaat acagtcatat 1261 gggactataa gcgggacgct ccggcgcata tatctaccat cggagtgtgc tcaatgacag 1321 atattgccaa gaaacctaca gagacgattt gcgctcctct taccgtattt tttgacggca 1381 gggtcgacgg acaagtggat ctgttccgaa acgcgcgcaa tggtgttctc atcactgagg 1441 gtagtgtaaa gggactccag cctagtgtcg gacctaagca agcctctttg aatggggtca 1501 ctctgatcgg cgaggctgtt aagacgcaat ttaactacta taagaaggtc gatggcgtcg 1561 tgcagcagct tcctgaaaca tactttacac agtctcgcaa cttgcaagaa tttaaaccgc 1621 gctcacagat ggaaatagac tttctcgaac tggcaatgga cgagttcata gagcggtaca 1681 aattggaagg gtatgccttt gaacacattg tttatggtga cttttctcac agccagttgg 1741 gtggtttgca tctcctcata gggctggcca aaaggttcaa agagagcccc tttgaattgg 1801 aggacttcat acctatggat tcaacagtta agaattactt cattacggat gcgcagacag 1861 gcagtagtaa gtgtgtctgc tccgtaattg acctcctctt ggacgacttt gtagagataa 1921 ttaagtccca ggatcttagt gtcgtatcca aggtcgttaa ggtcacaatt gattacaccg 1981 agattagttt tatgctgtgg tgtaaagacg gtcacgtcga gaccttctat ccaaagttgc 2041 aataaggatc caccggatct agataactga tcataatcag ccataccaca tttgtagagg 2101 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg 2161 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca 2221 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac 2281 tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa attcgcgtta 2341 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat 2401 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca 2461 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc 2521 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta 2581 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg 2641 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg 2701 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca 2761 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat 2821 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa 2881 aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa 2941 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa 3001 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca 3061 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca 3121 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg 3181 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct 3241 tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga 3301 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa 3361 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt 3421 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg 3481 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa 3541 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac 3601 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt 3661 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact 3721 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg 3781 ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg 3841 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc 3901 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt 3961 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc 4021 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg 4081 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg 4141 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct 4201 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc 4261 taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa 4321 gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg 4381 gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt 4441 ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg 4501 gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac 4561 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa 4621 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat 4681 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc 4741 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg 4801 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc 4861 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg 4921 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg 4981 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa 5041 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg 5101 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga 5161 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct 5221 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca 5281 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc 5341 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcattag ttattaatag 5401 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt 5461 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg 5521 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat 5581 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct 5641 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg 5701 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat g //