cleancap egfp mrna  (TriLink)

Journal: Molecular Therapy. Nucleic Acids

Article Title: mRNA-based in vivo induction of ETV2 to restore blood flow in a preclinical mouse hindlimb ischemia model

doi: 10.1016/j.omtn.2025.102592

Figure Lengend Snippet: Synthetic mRNA encoding ETV2 is efficiently translated into functional protein in vitro (A) EGFP or ETV2 protein expression at 15 h after mRNA transfection with MessengerMAX. Representative blight filed image and fluorescence image. Scale bars, 75 μm. TF, transfection reagent. (B) After mRNA transfection with MessengerMAX, the cells were collected at 24, 48, or 72 h time point. Total RNA extraction, cDNA synthesis, and qPCR analysis by ΔΔCT method were conducted. For each condition, n = 4; mean ± SD. ∗∗ p < 0.01 by two-way ANOVA with Bonferroni correction. (C) CDH5 protein expression at 24, 48, and 72 h time points after mRNA transfection were analyzed by western Blotting. The number above each gel lane represent the fold-change in intensity relative to control (TF only, 24 h).

Article Snippet: CleanCap EGFP mRNA, CleanCap Fluc mRNA, CleanCap Cre mRNA, and CleanCap Codon-optimized ETV2 mRNA (N1mΨ, RP-HPLC purified) were synthesized at TriLink Biotechnologies (San Diego, CA).

Techniques: Functional Assay, In Vitro, Expressing, Transfection, Fluorescence, RNA Extraction, cDNA Synthesis, Western Blot, Control

Journal: Molecular Therapy. Nucleic Acids

Article Title: mRNA-based in vivo induction of ETV2 to restore blood flow in a preclinical mouse hindlimb ischemia model

doi: 10.1016/j.omtn.2025.102592

Figure Lengend Snippet: In vivo biodistribution of the injected ETV2 mRNA with LNP and protein expression in skeletal muscle (A) ETV2 mRNA visualization with RNAscope. Quadriceps were collected 5 h after LNP (0.5 μg/limb) administration intramuscularly. Longitudinal section of the whole quadriceps was analyzed. Scare bars, 3 mm (low-magnification). (B and C) Representative images of (B) ETV2 mRNA visualization with RNAscope and (C) ETV2 protein visualization with IHC. Scale bars, 100 μm (high-magnification).

Article Snippet: CleanCap EGFP mRNA, CleanCap Fluc mRNA, CleanCap Cre mRNA, and CleanCap Codon-optimized ETV2 mRNA (N1mΨ, RP-HPLC purified) were synthesized at TriLink Biotechnologies (San Diego, CA).

Techniques: In Vivo, Injection, Expressing, RNAscope

Journal: Molecular Therapy. Nucleic Acids

Article Title: mRNA-based in vivo induction of ETV2 to restore blood flow in a preclinical mouse hindlimb ischemia model

doi: 10.1016/j.omtn.2025.102592

Figure Lengend Snippet: Profiling the cellular functional distribution of intramuscularly administrated mRNA/LNP (A) Schematic of the Ai6 transgenic mouse LoxP-flanked stop cassette preventing the transcription of ZsGreen. Upon delivery of Cre recombinase via Cre mRNA, the stop cassette is excised, and the cell expresses ZsGreen. (B) Experimental design of the functional delivery assay in vivo . (C) Gating strategy for each cell population: leukocytes, endothelial cells (ECs), and fibro/adipogenic progenitors (FAPs). (D) Flow cytometry scatter plots of ZsGreen expression in each cell population in PBS or Cre mRNA injected Ai6 mice. (E) ZsGreen expression in each cell population ( n = 2 mice). Data are represented as mean ± SD. ∗∗ p < 0.01.

Article Snippet: CleanCap EGFP mRNA, CleanCap Fluc mRNA, CleanCap Cre mRNA, and CleanCap Codon-optimized ETV2 mRNA (N1mΨ, RP-HPLC purified) were synthesized at TriLink Biotechnologies (San Diego, CA).

Techniques: Functional Assay, Transgenic Assay, In Vivo, Flow Cytometry, Expressing, Injection

Journal: Molecular Therapy. Nucleic Acids

Article Title: mRNA-based in vivo induction of ETV2 to restore blood flow in a preclinical mouse hindlimb ischemia model

doi: 10.1016/j.omtn.2025.102592

Figure Lengend Snippet: LNP encapsulating ETV2 mRNA promotes blood flow supply in a hindlimb ischemia model using BALB/c athymic mice (A) Experimental design of HLI study. On day 0 post the HLI surgery, PBS or LNP (3 μg/limb) was administered intramuscularly once a week up to 4 weeks. ETV2 lentivirus was administered intramuscularly on day 0 post HLI surgery. Blood flow was analyzed by LDI by day 35. (B) Representative LDI images on day 0 pre/post HLI surgery and day 14 post HLI surgery. (C) The ratio of blood flow (left injured limb/right normal limb) at all time points for all groups was shown. For each group, n = 6–15; mean ± SE. (D) The ratio of blood flow (left injured limb/right normal limb) at day 14 and day 28 was shown. ∗ p < 0.5 and ∗∗ p < 0.01 by two-way ANOVA with Bonferroni correction.

Article Snippet: CleanCap EGFP mRNA, CleanCap Fluc mRNA, CleanCap Cre mRNA, and CleanCap Codon-optimized ETV2 mRNA (N1mΨ, RP-HPLC purified) were synthesized at TriLink Biotechnologies (San Diego, CA).

Techniques:

Journal: Nature Communications

Article Title: Premixing enables loading of long RNA in cubic phase lipid nanoparticles

doi: 10.1038/s41467-025-60380-6

Figure Lengend Snippet: a – d Structural stability of the cubosome–long RNA complex. The SAXS study at various storage period ( a ) and temperature ( b ) shows no structural changes. In addition, no changes in the size of Q ll P, RNA ( c ) or leakage of long RNA from Q ll P, RNA ( d ) were found, indicating the excellent physical stability of the cubosome–long RNA complex. e Cell viability test of cubosome–long RNA complexes at 0.5 µg/mL concentration. The average value of the control group is set to 100%. Data are shown as mean values with SE and all data were obtained from three biologically independent experiments ( n = 3). f Evaluation of the cellular delivery efficiency of the cubosome–long RNA complexes. Cubosome–long RNA complexes were more effective than LNPs in delivering 1 kb. Data are shown as mean value (mean ± SE, n = 3; biological independent experiments). g The delivery performance was consistent even after storage at RT for 24 days (mean ± SE, n = 3; biological independent experiments). Statistical analysis was performed using ordinary one-way ANOVA followed by Tukey’s multiple comparisons test; no significant differences were observed ( p = 0.9798). h Visualization of the cubosome–long RNA complex and LNP in transfected HepG2 cells after 24 h. Lipids and mRNAs are shown in green (NBD-PE) and red (Cy5), respectively. Scale bar 25 µm. i , j Fluorescence intensites of cells containing lipids ( i ) and mRNAs ( j ) were quantified from ( h ) ( n = 3; individual cells pooled from independent replicates). k Protein expression image of cubosome-eGFP RNA complex transfection at 24 h post-treatment in HeLa cells. Green fluorescent image and merged with the bright field image are shown respectively. Scale bar 200 µm. l , m , Total fluorescence intensity of cells ( l ) and percentage of GFP expressing cells ( m ) were quantified from ( k ). All data are presented as mean ± SE, n = 3. Statistical analysis was performed using ordinary one-way ANOVA and Tukey’s multiple comparisons test (** p = 0.0041).

Article Snippet: CleanCap® EGFP mRNA (5moU) was customized from TriLink, and other kinds of RNA were purchased from Miltenyi Biotec and Sigma-Aldrich.

Techniques: Concentration Assay, Control, Transfection, Fluorescence, Expressing