vitro transcription  (New England Biolabs)

Journal: Synthetic and Systems Biotechnology

Article Title: Metabolic reprogramming of abscisic acid-producing strain Botrytis cinerea TB-31 toward terpenoid biosynthesis using a CRISPR/Cas9 ribonucleoprotein system

doi: 10.1016/j.synbio.2025.12.002

Figure Lengend Snippet: Transcriptomic analysis of B. cinerea strains TB-31, △ bcaba3 , and △ bcaba1234 . (A)∼(B) Regression analysis between transcriptomic data (RNA-seq) and RT‒qPCR validation. △ bcaba3 vs. TB-31: R 2 = 0.94, P < 0.001 (A); △ bcaba1234 vs. TB-31: R 2 = 0.97, P < 0.001 (B). (C) Heatmap of the relative transcriptional levels of the genes in the MVA pathway in △ bcaba1234 compared with those in TB-31. Gene transcription levels were normalized using Z score transformation, calculated as (X-mean)/SD, to represent the number of standard deviations above or below the mean transcription level. Upregulated genes are highlighted in red, while downregulated genes are shown in blue. The data are derived from three independent biological replicates. (D) Heatmap of the relative transcriptional levels of DEGs involved in other secondary metabolism pathways in △ bcaba1234 compared with those in TB-31. (E) Heatmap of the relative transcriptional levels of genes involved in intracellular primary metabolic pathways in the △ bcaba1234 strain compared with those in the TB-31 strain. Metabolic pathways are marked with gray boxes. PPP: pentose phosphate pathway; TCA: tricarboxylic acid cycle; GYC: glyoxylate cycle.

Article Snippet: The sgRNAs were transcribed using the T7 High Yield RNA Transcription Kit (Nanjing Vazyme Biotech Co., Ltd., Nanjing, China), and then purified using the RNA Clean & Concentrator Kit (Zymo Research Corp., Irvine, California, USA).

Techniques: RNA Sequencing, Biomarker Discovery, Transformation Assay, Derivative Assay

Journal: Transboundary and Emerging Diseases

Article Title: A Self‐Amplifying RNA Lipid Nanoparticle (saRNA‐LNP) Vaccine Provides Effective Protection Against Porcine Epidemic Diarrhea

doi: 10.1155/tbed/3115893

Figure Lengend Snippet: Production and characteristics of PEDV‐S mRNA and saRNA vaccines. (a) PEDV‐S saRNA vaccine design schematic. (b) Average particle size and PDI of RNA‐LNPs by dynamic light scattering. (c) qRT‐PCR quantification of the transcription efficiency in 293T cells transfected with saRNA‐ or mRNA‐LNPs at indicated time points. (d) Western blotting analysis PEDV‐S protein expression in 293T cells transfected with saRNA‐ or mRNA‐LNPs at indicated time points. (e) Densitometric quantification of PEDV‐S expression relative to β‐tubulin based on the blots of (d). Results in panels (c) and (e) were shown as mean ± SD of three independent experiments.

Article Snippet: The transcription reaction consisted of linearized template DNA, N1‐methylpseudouridine‐5 ′ ‐triphosphate (Novoprotein, China), CAP GAU m7G(5 ′ )ppp(5")(2 ′ 0MeA)pU (SYNTHGENE, China), and components from the T7 High‐Yield RNA Transcription kit (Novoprotein, China).

Techniques: Vaccines, Quantitative RT-PCR, Transfection, Western Blot, Expressing