LOCUS Exported 6512 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION mammalian expression and localization. ACCESSION . VERSION . KEYWORDS Nsp13-mCherry SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6512) 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 6512) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6512 /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 231..240 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" misc_feature 2043..2095 /label=MCS /note="multiple cloning site" regulatory 2097..2106 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2103..2813 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(2248..2266) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 2516..2535 /label=mCherry-F /note="mCherry, forward primer" polyA_signal 2936..3057 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2973..2992) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 3027..3046 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(3064..3519) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3201..3222) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3413..3432 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3546..3650 /gene="bla" /label=AmpR promoter promoter 3652..4009 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3675..3695) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3860..3995 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3922..3941 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 4044..4838 /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(4098..4117) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4708..4727 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(5026..5045) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 5070..5117 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 5446..6034 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5935..5954 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 6209..6512 /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 gccaccatgg 241 ctgttggggc gtgtgtgttg tgcaacagcc aaacctctct gcgatgcgga gcgtgcatac 301 ggaggccatt tctttgttgt aaatgttgtt atgatcacgt tattagtacg agccataagt 361 tggttctgag cgtcaacccc tatgtgtgta acgctccggg ttgcgacgta accgatgtga 421 cacaacttta cctggggggc atgtcatatt actgcaaaag tcacaaaccc ccaatctcat 481 ttccgttgtg tgcgaatggc caggtctttg gcctttacaa aaatacttgt gttggttcag 541 acaacgtaac cgattttaac gcgattgcta catgcgattg gacaaatgct ggtgattata 601 ttttggctaa tacgtgcacg gaaaggctca aattgtttgc agcggagacc ctcaaggcga 661 ccgaggaaac ctttaaactg agctacggca ttgcgacggt cagagaggtt ctttccgacc 721 gagaattgca cctctcatgg gaggtcggta agccgcgccc gcctctgaat aggaactatg 781 tctttacagg ctatcgagtt acaaaaaaca gtaaggtgca gataggggaa tatacttttg 841 agaaaggtga ctatggggac gccgtagtat acagggggac aactacctac aaactgaatg 901 tgggcgatta tttcgtgttg actagccata ctgtcatgcc cctctctgcc ccgaccctcg 961 ttccacagga acattatgtt cgcattactg ggctgtaccc cacgctcaac atatccgatg 1021 aatttagctc caacgtggca aactaccaga aggtgggaat gcaaaagtat agcacgttgc 1081 aaggcccccc tggcacaggt aaatcccact tcgccattgg cttggcgttg tactacccga 1141 gtgcccggat cgtctatact gcatgctctc atgctgcggt agacgcattg tgtgagaaag 1201 ccctgaagta tctgcccatt gacaagtgca gtcggataat cccagctcgg gcgcgagttg 1261 aatgcttcga taagtttaag gtaaatagta cacttgagca gtatgtattt tgtaccgtca 1321 acgctctccc cgaaaccact gcggacatag tggtatttga tgagatatct atggctacca 1381 attatgacct ctcagtagtc aacgcacgct tgcgagcaaa acactatgta tacatcggag 1441 acccagcgca attgcctgct cctaggactc tcctcaccaa agggactctt gagcctgaat 1501 atttcaactc cgtctgcagg ctgatgaaaa ccatcggacc cgatatgttt ctcgggacat 1561 gtcgaagatg tcctgccgaa attgtagata ctgttagcgc cttggtgtac gataataagc 1621 tgaaggccca taaagacaag agcgcccagt gtttcaagat gttctataag ggagtaatca 1681 cacatgacgt tagttctgcc atcaaccgac cgcagatagg cgtggtccgc gagttcctga 1741 cccgaaatcc agcgtggagg aaggccgtat ttatatcacc ttacaatagc caaaatgccg 1801 tggcgtcaaa gattttgggg ctcccgactc agacggtgga tagctctcaa gggtcagaat 1861 acgattacgt tatattcaca cagacgacag agacggcaca cagttgcaac gtcaacagat 1921 ttaatgtcgc tataacccgc gccaaggttg ggattctttg tataatgtct gacagagacc 1981 tctacgacaa gctgcagttc acgtcacttg agattcctcg caggaacgtt gcgacgcttc 2041 aactcgagct caagcttcga attctgcagt cgacggtacc gcgggcccgg gatcccgcca 2101 ccatggtgag caagggcgag gaggataaca tggccatcat caaggagttc atgcgcttca 2161 aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg 2221 gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt ggccccctgc 2281 ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc tacgtgaagc 2341 accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc aagtgggagc 2401 gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg 2461 acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccccg 2521 taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac cccgaggacg 2581 gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc cactacgacg 2641 ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacaacg 2701 tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg 2761 aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag taaagcggcc 2821 gcgactctag atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa 2881 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt 2941 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa 3001 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaagg 3061 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct 3121 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg 3181 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact 3241 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac 3301 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga 3361 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga 3421 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca 3481 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa 3541 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca 3601 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg 3661 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc 3721 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc 3781 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt 3841 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc 3901 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct 3961 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag 4021 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg 4081 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg 4141 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc 4201 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga 4261 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc 4321 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag 4381 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat 4441 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg 4501 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca 4561 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct 4621 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg 4681 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg 4741 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc 4801 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat 4861 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta 4921 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg 4981 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga 5041 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg 5101 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac 5161 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc 5221 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat 5281 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag 5341 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc 5401 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt 5461 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt 5521 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat 5581 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 5641 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 5701 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 5761 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 5821 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 5881 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa 5941 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt 6001 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg 6061 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc 6121 tgtggataac cgtattaccg ccatgcatta gttattaata gtaatcaatt acggggtcat 6181 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg 6241 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa 6301 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact 6361 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta 6421 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt 6481 acatctacgt attagtcatc gctattacca tg //