LOCUS Exported 5001 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION mammalian expression and localization. ACCESSION . VERSION . KEYWORDS EGFP-ORF9b SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5001) 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 5001) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5001 /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 1425..1546 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1462..1481) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1516..1535 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(1553..2008) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(1690..1711) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 1902..1921 /label=F1ori-R /note="F1 origin, reverse primer" promoter 2035..2139 /gene="bla" /label=AmpR promoter promoter 2141..2498 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(2164..2184) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 2349..2484 /label=SV40 ori /note="SV40 origin of replication" primer_bind 2411..2430 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 2533..3327 /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(2587..2606) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 3197..3216 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(3515..3534) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 3559..3606 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 3935..4523 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4424..4443 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 4698..5001 /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 tatggacccc aaaatttctg 1021 aaatgcaccc ggcgcttcgc ttggtggacc cccaaatcca actcgctgtc acgcggatgg 1081 aaaacgcagt aggacgcgac caaaataatg tcggcccaaa agtgtatccg attatcctcc 1141 gccttggcag tcccctgagc ctgaatatgg ccagaaagac tcttaacagc ctggaggaca 1201 aagcgtttca gctgaccccc attgcggttc agatgactaa gctggcgaca actgaagagt 1261 tgcccgacga atttgtggta gtaaccgtaa agtaaggatc caccggatct agataactga 1321 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc 1381 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag 1441 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt 1501 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc gtaaattgta 1561 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac 1621 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg 1681 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa 1741 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt 1801 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt 1861 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga 1921 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc 1981 gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga 2041 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa 2101 ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga aagaaccagc 2161 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta 2221 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag 2281 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa 2341 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac 2401 taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta ttccagaagt 2461 agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga gacaggatga 2521 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg 2581 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg 2641 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc 2701 ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct 2761 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa 2821 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg 2881 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa 2941 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat 3001 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg 3061 agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc 3121 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac 3181 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg 3241 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc 3301 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag 3361 cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg 3421 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc 3481 tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag acaataccgg 3541 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt gttgggtcgt 3601 ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc ccaccgagac 3661 cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc ccaagttcgg 3721 gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata gcctcaggtt 3781 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga 3841 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag 3901 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa 3961 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag 4021 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg 4081 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat 4141 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta 4201 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg 4261 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc 4321 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa 4381 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc 4441 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt 4501 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct 4561 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc 4621 gtattaccgc catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc 4681 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc 4741 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg 4801 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat 4861 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc 4921 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta 4981 ttagtcatcg ctattaccat g //