Solid Phase Reversible Immobilization (SPRI) is a technique applied in high-performance isolation, purification, and cleanup procedures. The process uses SPRI magnetic beads by selectively binding nucleic acids (involving RNA and DNA) found in all organisms.Solid Phase Reversible Immobilization (SPRI) is a technique applied in high-performance isolation, purification, and cleanup procedures. The process uses SPRI magnetic beads by selectively binding nucleic acids (involving RNA and DNA) found in all organisms. Researchers discovered SPRI in 1995 at the Whitehouse Institute for the isolation of PCR products. Specifically, the experiment focused on purifying PCR amplified colonies in the DNA sequencing group. SPRI beads play a significant role in next-generation sequencing (NGS) procedures. The SPRI technology enables researchers to concentrate on an amplified PCR (polymerase chain reaction) product like the ones used in COVID-19 tests. Additionally, SPRI eliminates excess reagents and nucleotides that cause inaccurate sequence data.
Characteristics and Chemical Reactions with SPRI BeadsHow does magnetic bead DNA extraction work? SPRI beads usually comprise a polystyrene core surrounded by magnetite (Fe3O4), which causes the beads to clump when exposed to a magnetic field. The surface of SPRI beads comprises carbonyl molecules that facilitate DNA binding. SPRI beads interact and activate with polyethylene glycol (PEG) and salt, and they provide a reversible binding process. The SPRI purification usually involves five general steps:
- Researchers bind and mix preparations to determine the size of DNA that binds in the experiment.
- A powerful magnet lumps the SPRI beads onto the DNA fragment targeted for purification.
- Researchers rinse the preparation with an alcohol (usually ethanol) solution.
- Scientists add a selective buffer to the preparation, which elutes DNA from the paramagnetic beads bound to the magnet.
- Researchers achieve DNA extraction by magnetic beads and remove a sample of the purified DNA for analysis and further testing.
Uses and Research with SPRI Beads
Identifying Plastic-associated SpeciesPlastic pollution remains a pressing global issue as more waste gets dumped into bodies of water. Scientists applied SPRI beads in identifying species in the Mediterranean Sea that arise from plastic pollution and its synthetic habitats. Researchers used DNA metacoding (using a small section of genes) to distinguish the taxonomic composition of plastic-based ocean populations. Researchers used PCR amplification for five barcode regions, which spanned a wide range of marine-plastic relevant species. They created sequencing libraries through two ligation steps, followed by DNA purification techniques using SPRI magnetic beads. The purification process enabled scientists to distinguish plastic-specific bacteria from naturally occurring water-born variants. Specifically, researchers compared species from seawater and polyethylene bags submerged for a month in two approximal underwater locations in the Mediterranean Sea. Results from the experiment identified 27 eukaryotic and 34 bacterial species unique to plastic habitats, of which 16 had associations with hydrocarbon degradation. The investigation suggested an efficient approach to characterizing the plastisphere and identifying its related species, which scientists may apply in other settings.
Plasmid PurificationResearchers applied SPRI in yielding high throughput DNA sequencing. SPRI was utilized to isolate and purify double-stranded plasmid DNA clones to support the capabilities of capillary electrophoresis DNA sequencing machines. Plasmids are DNA structures known for independent replication and are mostly studied and used in genetic engineering to amplify or copy specific genes. According to the experiment, SPRI proved effective as a polishing procedure through its scalability and automatability that purifies and prepares plasmid for DNA sequencing. Results showed an average increase of over 130 bases in read length (bases sequenced from a DNA fragment) and improved pass rate by ~20%.
Integrated Extraction of RNA and DNA From Viral PathogensAcute respiratory infections (ARIs) lead to severe diseases like pneumonia that associate with high mortality and morbidity rates. Researchers attributed the prevalence of ARIs to long turnaround times in diagnostics and inaccurate antibiotics prescriptions that lead to resistance and recovery complications. Conventional extraction methods collected RNA and DNA separately from sputum, leading to compounded time and cost factors in medical settings. Researchers applied SPRI in an experiment to establish a co-extraction method on a viral pathogen causing respiratory infections. The process examined the detection of two genera of respiratory viruses based on SPRI magnetic beads technology. Researchers evaluated influencing factors such as bead size, alcohol concentrations, lysis buffer pH, and RNA carrier type to determine the effectiveness of the co-extraction process. Results showed that the SPRI-based co-extraction method met the requirements of a multiple pathogen assay. Additionally, researchers observed that magnetic beads with carrier RNA significantly improved extraction efficiency by supporting nucleic acid precipitation.
ImmunoprecipitationImmunoprecipitation is a technique used to precipitate protein antigens out of a solution through a specific antibody that binds with it. The purification process enables researchers to perform various functions, such as measuring the molecular weight and identifying the activation status of proteins. Scientists have used SPRI magnetic beads as alternatives to agarose beads in immunoprecipitation and similar antibody precipitation processes. Experts prefer SPRI magnetic beads for multiple reasons, including reduced sample loss (eliminates centrifugation), faster separation process, and superior binding capacity (a surface area that optimizes antibody binding).
Purification of Dye-labeled DNA Sequencing FragmentsScientists have also applied SPRI through photoactivated polycarbonate (PC) to purify dye-labeled terminator sequencing fragments before electrophoretic gel sorting. Results showed that the PC-SPRI cleanup format effectively removed excess dye terminators but resulted in lower plate numbers. The experiment suggested improved SPRI efficiency through conventional off-chip preparations (using paramagnetic beads).
Improved Protocols in Illumina SequenceScientists commonly apply the Illumina sequence to compare DNA for an improved understanding of health and pathology. Illumina functions by breaking down DNA into manageable fragments and fuses with adaptor DNAs into a single strand with sodium hydroxide. Researchers applied SPRI toward improving cleanup and DNA-sizing protocols in Illumina sequencing. Results showed greater scalability in SPRI cleanup compared to standard column-based methods. Additionally, researchers noted that SPRI enabled the removal of most adapter dimers (a byproduct of PCR) after ligation. Similarly, researchers discovered advantages in DNA size selection with SPRI. The process enabled scientists to adjust preparations according to test requirements by setting bead to DNA ratios or shearing conditions for the desired DNA size range.
Advantages of HigherPurity™ PCR Clean-up Magnetic BeadsThe HigherPurity™ PCR Clean-Up Magnetic Beads system is a versatile and easily automated PCR system based on SPRI technology for PCR amplicon purification. These beads work effectively for various research, including cloning, fragment analysis, and sequencing. SPRI beads provide researchers with a long list of uses, such as removing excess salt and nucleotides with a quick and efficient wash.
The product comes preformulated to selectively bind DNA for easy use without organic extraction, centrifugation, or filtration steps.
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