One-pot method for preparing DNA, RNA, and protein for multiomics analysis.

Area of Science:

• Biochemistry
• Molecular Biology
• Genomics

Background:

• Traditional multiomics studies require separate, labor-intensive, and costly sample preparation methods for DNA, RNA, and proteins.
• Existing methods are prone to sampling bias, limiting comprehensive molecular profiling.
• There is a need for an integrated workflow to analyze multiple molecular layers from a single biological sample.

Purpose of the Study:

• To develop and validate a novel method for simultaneous preparation of high-quality DNA, RNA, and protein/peptides from a single sample.
• To enable comparative analysis of genomes, transcriptomes, and proteomes.
• To demonstrate the utility of this integrated multiomics workflow in cancer cell line and tissue analysis.

Main Methods:

• A reversible protein tagging scheme to link proteins to a bead-based matrix.
• Nucleic acid precipitation and selective solubilization techniques.
• Integrated workflow for simultaneous DNA, RNA, and protein/peptide isolation.

Main Results:

• Successful preparation of sequencing-ready DNA and RNA, and either intact proteins or mass-spectrometry-ready peptides.
• Demonstrated utility in comparing genomes, transcriptomes, and proteomes of triple-negative breast cancer cell lines.
• Identification of distinct RNA-associated protein and protein-only pathways differentiating cell lines.
• Successful application of the workflow to mouse tissue samples (brain, liver, lung).

Conclusions:

• The developed method offers a streamlined, cost-effective, and less biased approach for multiomics studies.
• This integrated workflow facilitates a more comprehensive understanding of molecular mechanisms in complex biological systems.
• The method is applicable to both cell line and tissue-based multiomics investigations.