Journal: Antioxidants

Article Title: Physio-Biochemical and Transcriptomic Features of Arbuscular Mycorrhizal Fungi Relieving Cadmium Stress in Wheat

doi: 10.3390/antiox11122390

Figure Lengend Snippet: RT-qPCR analysis of eight representative genes in the metabolic pathways of ‘glutathione metabolism’, ‘cysteine and methionine metabolism’, ‘plant hormone signal transduction’, and ‘metal ion transport’.

Article Snippet: Leaves were ground in liquid nitrogen, total RNA was isolated by the TRIzol reagent, and reverse transcription reactions were conducted by using PrimeScript ® RT Reagent Kit with gDNA Eraser (Takara, Kyoto, Japan). qRT-PCR experiments were performed on an ABI StepOne real-time PCR instrument by using Novozymes AceQ ® qPCR SYBR Green Master MIX kit.

Techniques: Quantitative RT-PCR, Transduction

Journal: bioRxiv

Article Title: Co-infection of influenza A virus enhances SARS-CoV-2 infectivity

doi: 10.1101/2020.10.14.335893

Figure Lengend Snippet: ( A and B ) A549 cells were mock-infected or infected with WSN at MOI 0.1. At 12 hours post-infection (h.p.i.), total RNAs were extracted from cells, and mRNA of ACE2, TMPRSS2, Furin, CatL ( A ), or mRNA of NP, Mx1, ISG54 ( B )was evaluated by quantitative real-time PCR (qRT-PCR) using SYBR green method. The data were expressed as fold changes relative to the Mock infections. ( C ) A549 cells were infected with WSN at MOI 0.1. IAV NP proteins (red) and ACE2 (green) were detected by an immunofluorescence assay using a confocal microscope at 12 hours-post-infection. Scale bars were shown. A549 ( D ), Calu-3 ( E ), and NHBE ( F ) cells were pre-infected with WSN at MOI 0.1 for 12 hours. Cells were then infected with live SARS-CoV-2 at MOI 0.01 for another 48 hours. Total RNAs were extracted from cells and mRNA of ACE2 was evaluated by quantitative real-time PCR (qRT-PCR) using SYBR green method. The protein expression levels of ACE2, SARS-CoV-2 N gene, IAV NP, and β-actin were measured by western blot. ( G ) The relative mRNA levels of ACE2 were measured from lung homogenates in indicated groups and the protein expression of IAV NP and ACE2 were detected by western blot accordingly. ( D-G and J ) The data were expressed as fold changes relative to the non-IAV infection control. ( H-K ) To establish ACE2 knock-down cells, A549 cell mixture was transduced with lentivirus encoding CRISPR-Cas9 system with two guide RNAs targeting ACE2 (sgRNA1 and sgRNA2) or control guide RNA respectively. Cells were infected with live SARS-CoV-2 at MOI 0.01 with or without IAV infection under the same procedure as above. The mRNA levels of ACE2 (qRT-PCR) ( H ) and SARS-CoV-2 E gene (Taqman-qRT-PCR) ( I ) expression were detected. ( J ) The mRNA level of ACE2 was detected by qRT-PCR in live SARS-CoV-2-infected cells transfected with either vector of WSN segment-2 respectively. ( K ) The mRNA levels of the SARS-CoV-2 E gene from either vector- or segment2-transfected cells were measured by Taqman-qRT-PCR at 48 hours post-live-SARS-CoV-2-infection in the present of control sgRNA or ACE2 sgRNAs. The data were expressed as fold change relative to non-IAV infection control. Values are mean ± s.d. of three independent results. *P≤0.05, **P≤0.01, ***P≤0.001, ****P≤0.0001.

Article Snippet: The primary antibodies used in this study were rabbit polyclonal antibody against ACE2 for immunofluorescence (Sino Biological, 10108-T26) and anti-influenza virus-NP (kindly provided by Prof. Ningshao Xia).

Techniques: Infection, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, SYBR Green Assay, Immunofluorescence, Microscopy, Expressing, Western Blot, Transduction, CRISPR, Transfection, Plasmid Preparation

Journal: bioRxiv

Article Title: Co-infection of influenza A virus enhances SARS-CoV-2 infectivity

doi: 10.1101/2020.10.14.335893

Figure Lengend Snippet: Calu-3 cells were mock-infected or infected with WSN at MOI of 0.1. At 12 hours h.p.i., total RNAs were extracted from cells, and mRNA of ACE2, TMPRSS2, Furin, CatL, NP, Mx1, and ISG54 were evaluated by qRT-PCR using the SYBR green method. The data were expressed as fold changes relative to the Mock infections. Values are mean ± s.d. of three independent results. *P≤0.05, **P≤0.01, ***P≤0.001, ****P≤0.0001.

Article Snippet: The primary antibodies used in this study were rabbit polyclonal antibody against ACE2 for immunofluorescence (Sino Biological, 10108-T26) and anti-influenza virus-NP (kindly provided by Prof. Ningshao Xia).

Techniques: Infection, Quantitative RT-PCR, SYBR Green Assay