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Rapid cloning-free mutagenesis of new SARS-CoV-2 variants using a novel reverse genetics platform.

Kipfer ET, Hauser D, Lett MJ, Otte F, Urda L, Zhang Y, Lang CMR, Chami M, Mittelholzer C, Klimkait T
Elife. 2023 Nov 21;12:RP89035. doi: 10.7554/eLife.89035. (Link opens in a new window) PubMed (Link opens in a new window) Article

Plasmids from Article

ID Plasmid Purpose
211731pUC19_CoV-2_linkerLinker fragment for direct rescue of SARS-CoV-2, including 5' Promoter and 3' Termination signals and 100bp SARS-CoV-2 homology regions
253115pUC19_CoV-2_frD1Encodes SARS-CoV-2 Wuhan spike
253116pUC19_CoV-2_frD_S Omicron BA.1For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), Wuhan backbone with Omicron BA.1 spike; chimeric
253117pUC19_CoV-2_frD_S Omicron BA.2For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), Wuhan backbone with Omicron BA.2 spike; chimeric
253119pUC19_CoV-2_frD1_XBB.1.5 SFor recombinant virus rescue; encodes fragment D1 of SARS-CoV-2 (nt20950-25522), Omicron isolate XBB.1.5 Spike
253121pUC19_CoV-2_frD_ΔE/GFP Δ68For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of envelope (residual AA) , orf6, orf8. GFP is inserted in the locus of envelope
253122pUC19_CoV-2_frD2_XBB_ΔE67 3*For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a, premature stop codon in orf3a at AA213. GFP is inserted in the locus of env
253124pUC19_CoV-2_frD_ΔORF3aFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of orf3a
253125pUC19_CoV-2_frD2_ΔORF6For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of orf6
253126pUC19_CoV-2_frD2_ΔORF7aFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of orf7a
253127pUC19_CoV-2_frD2_ΔORF7bFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of orf7b
253128pUC19_CoV-2_frD2_ΔORF8For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of orf8
253129pUC19_CoV-2_frD2_ΔORF68For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of orf6 and orf8
253130pUC19_CoV-2_frD2_ΔORF67a8For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of orf6, orf7a and orf8
253132pUC19_CoV-2_frD_ORF6-BirAFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), BirA fused to orf6, C-terminal
253144pUC19_CoV-2_frD ΔS/GFPFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of spike, GFP is inserted at the locus of Spike
253146pUC19_CoV-2_frD2 ΔE7 3* Δ6/H1 WSNFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; Influenza H1 (WSN) replaces orf6
253147pUC19_CoV-2_frD2 ΔE7 3* Δ6/H1 PR8For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; Influenza H1 (PR8) replaces orf6
253150pUC19_CoV-2_frD2_ΔrE/N1 PR8 Δ67 3*For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope (residual AA), orf6, orf7a;stop codon at AA213 in orf3a; Influenza N1 (PR8) replaces env
253151pUC19_CoV-2_frD2 ΔE6 3* Δr7a/H1 PR8For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a (residual AA); stop codon at AA213 in orf3a; Influenza H1 (PR8) replaces orf7a
253152pUC19_CoV-2_frD2_ΔrE/N1 Δ6 3* Δ7a/H1 PR8Virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of env (residual AA), orf6, orf7a;stop codon at AA213 in orf3a; Influenza N1 (PR8) replaces envelope; H1 (PR8) replaces orf7a
253153pUC19_CoV-2_frD2 ΔE6 3* Δr7a/H1 WSNFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), deletion of env, orf6, orf7a (residual AA);stop codon at AA213 in orf3a; Influenza H1 (WSN) replaces orf7a
253154pUC19_CoV-2_frD2_ΔrE/N1 WSN Δ67 3*For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of env (residual AA), orf6, orf7a; stop codon at AA213 in orf3a; Influenza N1 (WSN) replaces envelope
253155pUC19_CoV-2_frD2_ΔrE/N1 Δ6 3* Δ7a/H1 WSNVirus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of env (residual AA), orf6, orf7a; stop codon at AA213 in orf3a;Influenza N1 (WSN) replaces env;H1 (WSN) replaces orf7a
253157pUC19_CoV-2_frD_ΔE/F DS-Cav1 ΔCT ΔS67 3*For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of spike, envelope, orf6, orf7a;stop codon at AA213 in orf3a; stabilized RSV F replaces envelope
253160pUC19_CoV-2_frD_ΔrE/H1 WSN ΔTM ΔS67 3*For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of spike, env (residual AA), orf6, orf7a;stop codon at AA213 in orf3a; Influenza H1 (WSN) ΔTM replaces env
253161pUC19_CoV-2_frD2 ΔE6 3* Δ7a/H1 WSN ΔTMFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; Influenza H1 (WSN) ΔTM replaces orf7a
253162pUC19_CoV-2_ frD2_ΔE67 3* NA-P2A-N PR8For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; Influenza N1 (WSN)-P2A-N
253165pUC19_CoV-2_ frD2 ΔE6 3* Δr7a/H1 PR8 ΔTMFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; Influenza H1 (PR8) ΔTM replaces orf7a
253166pUC19_CoV-2_ frD2 ΔE6 3* Δr7a/N1 PR8For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a (residual AA);stop codon at AA213 in orf3a; Influenza N1 (PR8) replaces orf7a
253168pUC19_CoV-2_ frD_Δ3E/H1 PR8 Δ67aFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of orf3a, envelope, orf6, orf7a; Influenza H1 (PR8) replaces orf3a and envelope
253169pUC19_CoV-2_ frD_Δ3E/N1 PR8 Δ67aFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of orf3a, envelope, orf6, orf7a; Influenza N1 (PR8) replaces orf3a and envelope
253170pUC19_CoV-2_ frD2_ΔE67 3* VSVG-P2A-NFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; VSV glycoprotein fused to N, P2A
253171pUC19_CoV-2_ frD2 ΔE6 3* Δr7a/VSVGFor recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a (residual AA);stop codon at AA213 in orf3a; VSV glycoprotein replaces orf7a
253172pUC19_CoV-2_ frD_Δ3E/VSVG Δ67aFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; VSV glycoprotein replaces orf3a and envelope
253176pUC19_CoV-2_frD ΔS_RABVG CT CoV-2 ΔE67 3*For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of spike, env, orf6, orf7a;stop codon at AA213 in orf3a; rabies glycoprotein (CT SARS-2) replaces spike
253177pUC19_CoV-2_frD_Δ3E_RABVG Δ67aFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of orf3a, envelope, orf6, orf7a; rabies glycoprotein replaces orf3a and envelope
253178pUC19_CoV-2_frD ΔrS_GP LCMV ΔE67 3*For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of spike (partial), envelope, orf6, orf7a;stop codon at AA213 in orf3a; LCMV glycoprotein replaces spike
253179pUC19_CoV-2_frD_Δ3E_GP LCMV Δ67aFor recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867), with deletion of orf3a, envelope, orf6, orf7a; LCMV glycoprotein replaces orf3a and envelope
253180pUC19_CoV-2_frD2_ΔrE/esxA Δ67 3*For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a; Mycobacterium tuberculosis esxA replaces envelope
253183pUC19_CoV-2_frBFor recombinant virus rescue; encodes fragment B of SARS-CoV-2 (nt8590-15107)
253184pUC19_CoV-2_frCFor recombinant virus rescue; encodes fragment C of SARS-CoV-2 (nt15100-20958)
253185pUC19_CoV-2_frD_SV40For recombinant virus rescue; encodes fragment D of SARS-CoV-2 (nt20950-29867)
253186pUC19_CoV-2_frD2_ΔE67 3*For recombinant virus rescue; encodes fragment D2 of SARS-CoV-2 (nt25422-29867), with deletion of envelope, orf6, orf7a;stop codon at AA213 in orf3a

Antibodies from Article