BLADE-R: streamlined RNA extraction for molecular diagnostics and high-throughput applications.

Area of Science:

• Molecular Biology
• Genomics
• Biotechnology

Background:

• Efficient nucleic acid extraction is vital for molecular research.
• Current methods face challenges in large-scale clinical applications like RNA sequencing and PCR.
• There is a need for faster, cost-effective, and scalable nucleic acid purification techniques.

Purpose of the Study:

• To introduce BLADE-R, a novel magnetic bead-based protocol for simplified nucleic acid extraction and purification.
• To evaluate the efficiency, cost-effectiveness, and scalability of BLADE-R compared to traditional methods.
• To demonstrate the suitability of BLADE-R for various sample types and high-throughput applications.

Main Methods:

• Developed BLADE-R, a magnetic bead-based protocol combining lysis and binding in one step.
• Utilized on-bead rinsing for nuclease-free separation of DNA and RNA.
• Assessed RNA quality and yield using Agilent TapeStation and RT-qPCR.
• Performed RNA sequencing to compare gene expression profiles.
• Adapted the protocol for 96-well plate format for high-throughput processing.

Main Results:

• BLADE-R demonstrated superior time and cost efficiency compared to the TRIzol protocol for HEK293T cells.
• RNA sequencing showed comparable results between BLADE-R and TRIzol regarding sequence quality and gene count variance.
• BLADE-R outperformed TRIzol in RNA extraction from frozen tissue and whole blood samples.
• The 96-well plate format enabled purification of 96 samples in less time than single-sample traditional extraction.
• Minimal well-to-well contamination was observed in high-throughput processing.

Conclusions:

• BLADE-R offers an efficient, cost-effective, and scalable solution for nucleic acid extraction and purification.
• The protocol is effective across various sample types, including challenging ones like frozen tissue and whole blood.
• BLADE-R is adaptable to both low- and high-throughput formats, making it suitable for limited-resource settings.
• This method simplifies sample preparation for clinical PCR and sequencing assays.