human ace2 orf cdna  (Sino Biological)

Journal: Microbiology Spectrum

Article Title: O-GlcNAcylation at S659 enhances SARS-CoV-2 spike protein stability and pseudoparticle packaging efficiency

doi: 10.1128/spectrum.00527-25

Figure Lengend Snippet: O-GlcNAcylation of S at S659 regulates pseudoviral packaging of SARS-CoV-2. ( A ) S-S659A does not affect the binding between S and Ace2. HEK293T cells were transfected with the plasmids expressing HA-ACE2 and GFP-S-WT, GFP-S-659A, or empty vector. The cell lysates were subject to IP. ( B ) Quantitation of ( A ). ( C ) Diagram of four-plasmid system for SARS-CoV-2pp production. The four-plasmid system for preparing pseudovirus SARS-CoV-2pp includes Luc (pLenti6) encoding luciferase, HIV Gag/pol (pLP1), HIV Rev (pLP2), and GFP-S. ( D ) S659A mutation in SARS-CoV-2 S reduced the SARS-CoV2pp package efficiency. The luciferase activity of lysates from Huh7.5.1 cells infected with SARS-CoV-2pp generated from HEK293T cells based on either S or the S-S659A was assayed. ns, no significance; ****, P < 0.0001.

Article Snippet: The human ACE2 ORF cDNA clone expression plasmid with a C-HA tag was sourced from Sino Biological (HG10108-CY).

Techniques: Binding Assay, Transfection, Expressing, Plasmid Preparation, Quantitation Assay, Luciferase, Mutagenesis, Activity Assay, Infection, Generated

Journal: Microbiology Spectrum

Article Title: O-GlcNAcylation at S659 enhances SARS-CoV-2 spike protein stability and pseudoparticle packaging efficiency

doi: 10.1128/spectrum.00527-25

Figure Lengend Snippet: The model of O-GlcNAcylation of the SARS-CoV-2 S protein. The S protein is modified by O-GlcNAcylation, which is promoted by OGT and reduced by OGA. O-GlcNAcylation enhances the stability of the S protein, while non-GlcNAcylated S is targeted for ubiquitination and degradation. Thiamet G, an OGA inhibitor, elevates the levels of O-GlcNAcylation on the S protein. Proper glycosylation of the S protein is essential for the entry of SARS-CoV-2pp, which is mediated by the ACE2 receptor.

Article Snippet: The human ACE2 ORF cDNA clone expression plasmid with a C-HA tag was sourced from Sino Biological (HG10108-CY).

Techniques: Modification, Ubiquitin Proteomics, Glycoproteomics

Journal: Frontiers in Microbiology

Article Title: Generation and characterization of a humanized ACE2 rat model for the study of SARS-CoV-2 and COVID-19

doi: 10.3389/fmicb.2025.1680273

Figure Lengend Snippet: Relative mRNA detection and western blot analysis of Human ACE2 expression. Kidney (K), lung (L) and brain (B) tissues were collected from the indicated transgenic hemizygous (HEM); homozygous (HOM) and wild type (WT) rats. (A,D) F344-Tg(CAG- ACE2 )057Bryd; (B,E) SD-Tg( ACE2 )955CPBryd; (C,F) SD-Tg( ACE2 )058CVBryd line. (A–C) RT-PCR analysis. The rat reference gene Hprt1 was used to normalize human ACE2 mRNA expression; each bar represents expression for an individual rat. Error bars represent the standard deviations between triplicate technical replicates. (D–F) Western blot analysis., HepG2 cells (PC) were used as a positive control and Rat Hprt was used as the loading control. M is the molecular size standard lane with the sizes of the standards indicated in kDa.

Article Snippet: The human ACE2 cDNA was purchased from Origene (Catalog #RG208442) and subcloned into pCAG-loxP-STOP-loxP-ZsGreen (Addgene plasmid #51269) using restriction enzymes Sal I and Sac I.

Techniques: Western Blot, Expressing, Transgenic Assay, Reverse Transcription Polymerase Chain Reaction, Positive Control, Control

Journal: Frontiers in Microbiology

Article Title: Generation and characterization of a humanized ACE2 rat model for the study of SARS-CoV-2 and COVID-19

doi: 10.3389/fmicb.2025.1680273

Figure Lengend Snippet: F344-Tg(CAG- ACE2 )057Bryd transgenic rats are susceptible to SARS-CoV-2 challenge. Groups of F344-Tg(CAG- ACE2 )057Bryd littermates without the ACE2 transgene (wild type, WT, orange circles), or hemizygous (HEMI, blue squares) or homozygous (HOM, purple triangles) for transgene expression were challenge with SARS-CoV-2 by intranasal instillation of 14 PFU of the USA-WA1/2020 variant. Rats were monitored for survival (A) and clinical symptoms including weight loss (B) for up to 10 days. Weight loss of hemizygous animals has been separated into those that survived (solid lines, n = 2) and those that succumbed to disease (dashed lines, n = 10). Animals identified as moribund were humanely euthanized, and tissues were collected for histological analysis (see , ). N = 7 (WT), 13 (HEMI), and 6 (HOM) collected in two independent experiments. Survival data was analyzed by Mantel-Cox (log-rank) test. Daily change in weight was analyzed by ANOVA with Dunn’s multiple comparison test. * P < 0.05, ** P < 0.01, *** P < 0.001 relative to WT. Survival of hemizygous and homozygous groups were not significantly different from each other.

Article Snippet: The human ACE2 cDNA was purchased from Origene (Catalog #RG208442) and subcloned into pCAG-loxP-STOP-loxP-ZsGreen (Addgene plasmid #51269) using restriction enzymes Sal I and Sac I.

Techniques: Transgenic Assay, Expressing, Variant Assay, Comparison

Journal: Frontiers in Microbiology

Article Title: Generation and characterization of a humanized ACE2 rat model for the study of SARS-CoV-2 and COVID-19

doi: 10.3389/fmicb.2025.1680273

Figure Lengend Snippet: Tissue damage of F344-Tg(CAG- ACE2 )057Bryd rats challenged with SARS-CoV-2. Sections of lung (A,D) , eye (B,E) and nasal cavity (C,F) from wild type (A–C) and F344-Tg(CAG- ACE 2)057Bryd hemizygous rats infected with either high (D) or low (E,F) doses of SARS-CoV-2 stained with hematoxylin and eosin. (D) One hemizygous rat infected with the high dose demonstrates broncho and interstitial pneumonia. Bronchiolar lumina are multifocally filled with mucinous material and low numbers of neutrophils; alveolar septae and lumina contain alveolar macrophages, lymphocytes and mild hemorrhage. (E) Hemizygous rats infected with the low dose show anterior uveitis characterized by infiltration of the iris and choroid body with neutrophils and lymphocytes and anterior chamber accumulations of eosinophilic flocculent material, neutrophils, few lymphocytes and red blood cells. The corneal stroma is infiltrated by moderate numbers of neutrophils. (F) Hemizygous rats infected with the low dose show foci of epithelial erosion and associated mucosal neutrophilic infiltrates as well as accumulation of suppurative exudate in the nasal cavity. All images taken at 200× magnification.

Article Snippet: The human ACE2 cDNA was purchased from Origene (Catalog #RG208442) and subcloned into pCAG-loxP-STOP-loxP-ZsGreen (Addgene plasmid #51269) using restriction enzymes Sal I and Sac I.

Techniques: Infection, Staining

Journal: Frontiers in Microbiology

Article Title: Generation and characterization of a humanized ACE2 rat model for the study of SARS-CoV-2 and COVID-19

doi: 10.3389/fmicb.2025.1680273

Figure Lengend Snippet: Cellular changes in the brains of F344-Tg(CAG- ACE2 )057Bryd rats that succumbed to a high dose SARS-CoV-2 challenge. Sections of cerebrum and cerebellum from wild type (A–D) and F344-Tg(CAG- ACE2 )057Bryd hemizygous rats (E–H) infected with SARS-CoV-2 stained with hematoxylin and eosin (A,D,E,H) or subjected to IBA (B,F) or GFAP (C,G) immunohistochemistry. (E) Histology of hemizygous rats exhibit macrophagic meningitis, perivascular cuffing, encephalitis and necrosis. (F) IBA IHC demonstrates marked glial cell activation. (G) GFAP IHC shows astrocyte proliferation swelling in areas of gliosis. (H) Cerebellum exhibits Purkinje cell loss associated with vacuolated neuropil of the adjacent molecular layer and scattered pyknotic and karyorrhectic debris (apoptosis as determined by PCNA IHC) with mildly increased numbers of astrocytes or microglia. All images taken at 200× magnification.

Article Snippet: The human ACE2 cDNA was purchased from Origene (Catalog #RG208442) and subcloned into pCAG-loxP-STOP-loxP-ZsGreen (Addgene plasmid #51269) using restriction enzymes Sal I and Sac I.

Techniques: Infection, Staining, Immunohistochemistry, Activation Assay