LOCUS Exported 6227 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION mammalian expression and localization. ACCESSION . VERSION . KEYWORDS Nsp13 deltaN1-EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6227) 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 6227) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6227 /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 1743..1795 /label=MCS /note="multiple cloning site" regulatory 1803..1812 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1809..2528 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 1809..2525 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1854..1875) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2115..2134) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2462..2483 /label=EGFP-C /note="EGFP, forward primer" polyA_signal 2651..2772 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2688..2707) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2742..2761 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2779..3234) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2916..2937) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3128..3147 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3261..3365 /gene="bla" /label=AmpR promoter promoter 3367..3724 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3390..3410) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3575..3710 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3637..3656 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3759..4553 /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(3813..3832) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4423..4442 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4741..4760) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4785..4832 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 5161..5749 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5650..5669 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 5924..6227 /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 acaacgtaac cgattttaac gcgattgcta catgcgattg gacaaatgct ggtgattata 301 ttttggctaa tacgtgcacg gaaaggctca aattgtttgc agcggagacc ctcaaggcga 361 ccgaggaaac ctttaaactg agctacggca ttgcgacggt cagagaggtt ctttccgacc 421 gagaattgca cctctcatgg gaggtcggta agccgcgccc gcctctgaat aggaactatg 481 tctttacagg ctatcgagtt acaaaaaaca gtaaggtgca gataggggaa tatacttttg 541 agaaaggtga ctatggggac gccgtagtat acagggggac aactacctac aaactgaatg 601 tgggcgatta tttcgtgttg actagccata ctgtcatgcc cctctctgcc ccgaccctcg 661 ttccacagga acattatgtt cgcattactg ggctgtaccc cacgctcaac atatccgatg 721 aatttagctc caacgtggca aactaccaga aggtgggaat gcaaaagtat agcacgttgc 781 aaggcccccc tggcacaggt aaatcccact tcgccattgg cttggcgttg tactacccga 841 gtgcccggat cgtctatact gcatgctctc atgctgcggt agacgcattg tgtgagaaag 901 ccctgaagta tctgcccatt gacaagtgca gtcggataat cccagctcgg gcgcgagttg 961 aatgcttcga taagtttaag gtaaatagta cacttgagca gtatgtattt tgtaccgtca 1021 acgctctccc cgaaaccact gcggacatag tggtatttga tgagatatct atggctacca 1081 attatgacct ctcagtagtc aacgcacgct tgcgagcaaa acactatgta tacatcggag 1141 acccagcgca attgcctgct cctaggactc tcctcaccaa agggactctt gagcctgaat 1201 atttcaactc cgtctgcagg ctgatgaaaa ccatcggacc cgatatgttt ctcgggacat 1261 gtcgaagatg tcctgccgaa attgtagata ctgttagcgc cttggtgtac gataataagc 1321 tgaaggccca taaagacaag agcgcccagt gtttcaagat gttctataag ggagtaatca 1381 cacatgacgt tagttctgcc atcaaccgac cgcagatagg cgtggtccgc gagttcctga 1441 cccgaaatcc agcgtggagg aaggccgtat ttatatcacc ttacaatagc caaaatgccg 1501 tggcgtcaaa gattttgggg ctcccgactc agacggtgga tagctctcaa gggtcagaat 1561 acgattacgt tatattcaca cagacgacag agacggcaca cagttgcaac gtcaacagat 1621 ttaatgtcgc tataacccgc gccaaggttg ggattctttg tataatgtct gacagagacc 1681 tctacgacaa gctgcagttc acgtcacttg agattcctcg caggaacgtt gcgacgcttc 1741 aactcgagct caagcttcga attctgcagt cgacggtacc gcgggcccgg gatccaccgg 1801 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg 1861 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg 1921 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct 1981 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc 2041 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca 2101 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg 2161 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc 2221 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc 2281 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc 2341 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg 2401 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc 2461 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt 2521 acaagtaaag cggccgcgac tctagatcat aatcagccat accacatttg tagaggtttt 2581 acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat 2641 tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac 2701 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat 2761 caatgtatct taaggcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt 2821 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat 2881 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat 2941 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac 3001 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc 3061 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga 3121 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca 3181 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg 3241 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa 3301 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga 3361 agagtcctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc 3421 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag 3481 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta 3541 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc 3601 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc 3661 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg 3721 caaagatcga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc 3781 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga 3841 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt 3901 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat 3961 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg 4021 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg 4081 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc 4141 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga 4201 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag 4261 ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc 4321 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg 4381 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata 4441 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg 4501 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac 4561 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc 4621 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat 4681 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac 4741 tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca 4801 gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg 4861 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt 4921 ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc 4981 ggcaggccct gccatagcct caggttactc atatatactt tagattgatt taaaacttca 5041 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc 5101 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc 5161 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc 5221 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt 5281 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt 5341 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc 5401 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa 5461 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac 5521 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg 5581 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga 5641 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact 5701 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa 5761 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc 5821 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat 5881 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg 5941 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt 6001 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac 6061 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg 6121 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact 6181 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatg //