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Single-Base tRNA Modification Sequencing
Single-Nucleotide tRNA Pseudouridine (Ψ) Modification Seq(BACS-seq)
BACS-seq is an antibody-free, base-resolution route that protects pseudouridine on tRNA. Controlled bisulfite conversion shields Ψ and reads it as a unique signature while other bases are changed, giving absolute base-resolution Ψ quantification.
Arraystar Single-Nucleotide tRNA Pseudouridine (Ψ) Modification Seq(BACS-seq) is an end-to-end sample-to-data service for tRNA, from RNA sample QC and tRNA treatment through library construction, sequencing, and bioinformatics. Sites are localized within mature tRNA transcripts with relative coordinates, tRNA gene attribution, anticodon and isotype, and structural-domain localization.
Benefits
In tRNA, pseudouridine stabilizes structure and decoding, so site-level mapping is informative. Controlled bisulfite conversion shields Ψ and reads it as a unique signature while other bases change, resolving each Ψ site at base resolution.
Single-base resolution: pinpoints Ψ sites in tRNA.
Quantitative measurement: the conversion read-out yields reliable modification fractions.
No antibody dependence: eliminates enrichment bias and cross-reactivity concerns.
Functional annotation: Sites are localized within mature tRNA transcripts with relative coordinates, tRNA gene attribution, anticodon and isotype, and structural-domain localization.
Application-ready: supports decoding fidelity, codon use, aminoacylation, and tRNA structural biology across tRNA isotypes.
| Service Name | Price |
|---|---|
| Single-Nucleotide tRNA Pseudouridine (Ψ) Modification Seq(BACS-seq) |
Background
In tRNA, the modification stabilizes the folded L-shaped structure and fine-tunes anticodon–codon decoding, so base-resolution mapping within maturation loops and the anticodon is what distinguishes it. Controlled bisulfite chemistry protects pseudouridine and reads it as a distinct signature, while unmodified positions are converted and read normally, giving absolute base-resolution pseudouridine quantification. This antibody-free readout maps each Ψ site at single-base resolution.
The method was validated for tRNA, sites are localized within mature tRNA transcripts with relative coordinates, tRNA gene attribution, anticodon and isotype, and structural-domain localization. (Xu H, et al. Song CX. Nature Methods 2024;21(11):2024-2033., DOI: 10.1038/s41592-024-02439-8).
Key concept: Single-Nucleotide tRNA Pseudouridine (Ψ) Modification Seq(BACS-seq) is a base-resolution method in which controlled bisulfite chemistry protects pseudouridine and reads it as a distinct signature while unmodified bases are converted, enabling absolute base-resolution Ψ quantification.
Figure 1. BACS-seq pseudouridine detection scheme. Selective bisulfite treatment shields Ψ and marks it as a unique signature while other bases are chemically converted, yielding absolute base-resolution quantification.
Workflow
A streamlined workflow turns the RNA sample into base-resolution pseudouridine (Ψ) datasets:
Figure 2. Single-Nucleotide tRNA Pseudouridine (Ψ) Modification Seq workflow.
Bioinformatics
Downstream analysis aligns reads to a curated reference, calls BACS-seq conversion signatures, and reports Ψ stoichiometry per position alongside motif, distribution, differential, enrichment, and genome-browser results.
Deliverables
- Raw Ψ tRNA sequencing FASTQ data files, formatted for GEO/SRA public database submission.
- QC report with BAM alignment files and key mapping metrics.
- High-confidence single-base Ψ tRNA site tables (XLSX) with coordinates, anticodon, isotype, and stoichiometry.
- Single-base Ψ distribution plots across tRNA domains and consensus Ψ motif logos (PDF/PNG).
- Single-base resolution differential Ψ tRNA site tables (XLSX) with fold change and significance.
- Genome browser-compatible Ψ signal track files (bigWig/bedGraph) and a structured tRNA report.
Research Applications
- tRNA decoding fidelity: maps the modification within anticodon and wobble regions.
- Codon use & aminoacylation: relates base-resolution modification to charging and translation.
- tRNA structural biology: localizes sites within D-loop, anticodon, TΨC, and acceptor stem.
- Isotype-resolved analysis: compares modification across tRNA isotypes and isoacceptors.
- Disease-oriented studies: finds differentially modified tRNA sites as biomarkers.
References
[1] Xu H, Kong L, Cheng J, et al. Song CX. Absolute quantitative and base-resolution sequencing reveals comprehensive landscape of pseudouridine across the human transcriptome. Nature Methods 2024;21(11):2024-2033. DOI: 10.1038/s41592-024-02439-8.
Sample Requirements
Sample Storage
For cells/tissue: use TRIzol or an RNA-stabilizing reagent, quick-freeze in liquid nitrogen, and keep at –80 °C.
For RNA: dissolve in ethanol or RNase-free water, store at –80 °C, and limit freeze–thaw cycles.
Dispatch Guidelines
Transfer each sample into a 1.5 mL nuclease-free tube.
Close the tube securely with parafilm or a cap lock to preserve integrity.
Send on dry ice with sufficient insulation to sustain the required temperature.
| Sample Type | Notes |
|---|---|
| Whole blood | Use EDTA tubes only, as heparin is not compatible with subsequent analytical procedures. |
| Cultured cells | Submission of cell pellets is preferred to ensure high-quality material for processing. |
| Tissue | Provide fresh or frozen specimens, ensuring that necrotic material is strictly avoided. |
| Total RNA | Maintain an OD 260/280 ratio ≥ 1.8 and RIN ≥ 7 with no visible degradation. Submit at least 10 µg total RNA. |
FAQ
Which tRNA features are annotated?
Sites are localized within mature tRNA transcripts with relative coordinates, tRNA gene attribution, anticodon and isotype, and structural-domain localization. This lets you interpret each site in the context of its host tRNA isotype and structural domain. The method supports both mRNA and tRNA workflows, so you can choose the readout that matches your study.
Does this method measure the modification at single-base resolution?
Yes. Controlled bisulfite chemistry protects pseudouridine and reads it as a distinct signature, while unmodified positions are converted and read normally, giving absolute base-resolution pseudouridine quantification. This provides single-base resolution and absolute stoichiometry for each site in tRNA.
How does this method differ from antibody-based tRNA profiling?
Antibody-based methods rely on enrichment and offer limited resolution with intrinsic motif bias. This method is antibody-free and delivers base-resolution, quantitative calling in tRNA, avoiding cross-reactivity and enrichment bias. This suits projects needing quantitative site-level mapping rather than region-level enrichment, and it pairs well with orthogonal validation.
How does this service support pseudouridine research?
By detecting Ψ with high accuracy, the service helps researchers study its functional significance, offering valuable insight for translational studies.
Why is accurate pseudouridine detection a valuable research asset?
Reliable, quantitative pseudouridine profiling strengthens the evidence base for research findings, making it a dependable tool for advancing the understanding of RNA function.
!This assay is provided for research applications only and is not intended for clinical diagnosis, treatment, or personal health evaluation.