A high-throughput pipeline for DNA/RNA/small RNA purification from tissue samples for sequencing.

Area of Science:

• Molecular Biology
• Biotechnology
• Genomics and Transcriptomics

Background:

• Standard solid-phase reversible immobilization (SPRI) bead protocols efficiently purify total nucleic acids (TNA) for genomic and transcriptomic analyses.
• However, these methods exhibit weak binding of small RNA species, such as microRNA (miRNA), necessitating separate, difficult-to-automate column-based purification workflows.

Purpose of the Study:

• To develop a single, high-throughput, SPRI bead-based protocol for the simultaneous purification of DNA, RNA, and small RNA.
• To enhance small RNA recovery within an automated, plate-based workflow for next-generation sequencing (NGS) applications.

Main Methods:

• A modified SPRI bead protocol, termed GSC-modified RLT+ Aline bead-based protocol (GRAB-ALL), was developed incorporating specific modifications to improve small RNA yield.
• The GRAB-ALL protocol was benchmarked against established nucleic acid purification workflows.

Main Results:

• GRAB-ALL demonstrated efficient purification of TNA, encompassing DNA, RNA, and crucially, small RNA species.
• The protocol is designed for a plate-based format, making it suitable for automated, high-throughput sample preparation.

Conclusions:

• The GRAB-ALL protocol offers an integrated solution for high-throughput TNA purification, significantly improving small RNA recovery compared to standard SPRI methods.
• This advancement facilitates streamlined sample preparation for diverse NGS applications, including those requiring small RNA analysis.