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Yes. Nsun2 rescue experiments are well-established for RNA modification research: • Construct design: use a codon-modified Nsun2 sequence with a small C-terminal tag (FLAG, HA). Nsun2 has N-terminal SAM-binding methyltransferase domain and C-terminal regulatory region — preserve both. • Catalytically-dead rescue: C271A/C321A double mutation in the catalytic cysteines abolishes methyltransferase activity and is the standard specificity control. • Functional readout: rescue should restore tRNA and mRNA m5C levels measured by bisulfite sequencing or mass spectrometry. TC1 mESCs require feeder cell co-culture or feeder-free conditions with LIF — lentiviral transduction of mESCs is supported but requires optimization; transgene silencing during differentiation should be monitored using silencing-resistant promoters. Confirm pluripotency markers (Oct4, Nanog, SSEA-1) and karyotype stability after rescue line generation.

Primary applications: • tRNA m5C analysis: bisulfite sequencing or mass spectrometry analysis of tRNA m5C levels (notably tRNA-Leu C34/C38, tRNA-Gly). • mRNA m5C analysis: m5C-RIP-seq or bisulfite-seq to characterize Nsun2-dependent mRNA m5C deposition. • Stem cell self-renewal: pluripotency marker (Oct4, Nanog) expression and self-renewal assays in mESC context. • Differentiation studies: directed differentiation of mESCs into the three germ layers to characterize m5C's role in lineage specification. EDITGENE recommends this mouse ESC-based model for researchers investigating m5C epitranscriptomics, RNA modification in pluripotency, and Nsun2-dependent stem cell biology.

The choice depends on whether you are studying Nsun2 (NOL1/NSUN2)'s role as the principal m5C tRNA methyltransferase or its emerging functions in mRNA m5C methylation and pluripotency. The Knockout line is the standard tool for asking whether Nsun2 is required for m5C deposition — Nsun2 generates 5-methylcytosine (m5C) modifications on tRNAs (notably tRNA-Leu C34 region), mRNAs, and ncRNAs, with established roles in stem cell biology and Dubowitz-like syndrome (NSUN2 mutations cause autosomal recessive intellectual disability). Overexpression is useful for studying Nsun2 in heterologous expression contexts. For RNA modification and stem cell research, the EDITGENE Nsun2 Knockout in TC1 mouse ESC is highly valuable — TC1 is a 129S6/SvEvTac-derived mouse embryonic stem cell line, providing a pluripotent stem cell background to study m5C's role in self-renewal and differentiation. Rescue with wild-type or catalytically-dead (C271A/C321A double mutation in the catalytic cysteines) Nsun2 enables structure-function studies. The mouse ESC background uniquely enables differentiation studies to characterize m5C's role in lineage specification. The model is valuable for m5C epitranscriptomics research.

Transfecting suspension cells is generally more challenging. However, due to its superior performance, this product works efficiently not only in adherent cells but also in suspension cells. For example, in Jurkat cells, 48 hours post-transfection, editing efficiency can reach up to 97%, demonstrating the product’s high efficiency in suspension cell transfection and meeting demanding requirements.

The product utilizes advanced biomolecular transfection technology. Compared with the toxicity of traditional chemical transfection methods and the physical stress of electroporation, it shows significant advantages in preserving cell viability.

If gene knockout fails when using this kit, EDITGENE will not charge for the kit. Additionally, the fee you paid for the kit can be directly applied toward EDITGENE’s gene knockout service, ensuring that your gene editing experiments proceed without concerns.

The product has been validated in multiple cell types. The RNP system enters cells and begins functioning within 4 hours post-transfection, and the Cas9 protein is degraded within 24-48 hours. This transient, high-efficiency expression enables gene editing without the need for continuous selection.

While suspension cells are generally more difficult to transfect, this kit performs exceptionally well in both adherent and suspension cell types. For example, in Jurkat cells, editing efficiency can reach up to 97% within 48 hours post-transfection, demonstrating the kit’s outstanding performance and suitability for demanding suspension cell applications.

This kit has been validated across multiple cell lines. The RNP complex enters cells and begins gene editing within 4 hours post-transfection. Cas9 protein is degraded within 24–48 hours, allowing efficient and transient expression-driven editing without the need for antibiotic or fluorescent selection.

The kit employs advanced biomolecular transfection technology, offering significant advantages over traditional methods. Unlike chemical transfection, which may be cytotoxic, or electroporation, which can subject cells to physical stress, this approach ensures minimal damage while maintaining high efficiency.

The kit can detect Mycoplasma in common cell culture reagents, such as media and serum, without the need for sample preparation. Simply proceed directly to PCR reaction and gel loading. However, the kit cannot detect Mycoplasma in organic solvents like DMSO or ethanol.

Use a freshly opened negative control for retesting. Ensure to change pipette tips during sample loading and prioritize loading the negative control first to avoid cross-contamination between samples.

PCR detection of Mycoplasma is highly sensitive, so avoid cross-contamination between samples. Ensure sterile conditions when handling each sample individually. After sample processing and PCR reaction, allow samples to cool before proceeding to the next step to avoid aerosol contamination.

The kit can detect Mycoplasma in common cell culture reagents, such as media and serum, without the need for sample preparation. Simply proceed directly to PCR reaction and gel loading. However, the kit cannot detect Mycoplasma in organic solvents like DMSO or ethanol.

PCR detection of Mycoplasma is highly sensitive, so avoid cross-contamination between samples. Ensure sterile conditions when handling each sample individually. After sample processing and PCR reaction, allow samples to cool before proceeding to the next step to avoid aerosol contamination.