An SPRI beads-based DNA purification strategy for flexibility and cost-effectiveness

Abstract

Background Current solid-phase reversible immobilization (SPRI) beads technology is widely used in molecular biology due to its convenience for DNA manipulation. However, the high performance commercial SPRI beads have no price advantage over our method. Furthermore, the use of commercially available SPRI beads standards does not provide the flexibility required for a number of specific nucleic acid handling scenarios. Results We report an efficient DNA purification strategy by combining home-made beads-suspension buffer with SPRI beads. The method tests the critical concentrations of polyethylene glycol (PEG) 8000 and beads to maximise recovery. And the composition of the SPRI beads DNA purification system (SDPS) was determined at 20% PEG 8000, 2 M NaCl and 16.3 mM MgCl2, and 1.25 mg/ml beads (1/8th original concentration). Then, we tested the DNA recovery of the SDPS, and the result showed that it was comparable to the control (AMPure XP beads). In the study, we have also developed an adjustment SPRI beads DNA purification system (ASDPS), the volume of ASDPS per reaction is 0.6× reaction volume (beads/samples). The performance of ASDPS is similar to SDPS and the control. But the cost of our methods is only about 1/24th of the control. To further assess its performance, we prepare the DNA-seq libraries to evaluate the yield, library quality, capture efficiency and consistency. We have compared all these results with the performance of the control and confirmed its efficiency. Conclusion We have proposed an alternative DNA purification approach with great flexibility, allowing researchers to manipulate DNA in different conditions. And ultimately, its application will benefit molecular biology research in the future.

Pneumatic nano-sieve for CRISPR-based detection of drug-resistant bacteria

“…In this process, magnet beads are introduced into a solution containing DNA, wherein a substantial amount of salt and polyethylene glycol (PEG) is present. 36 The DNA molecules become crowded out and bind to the surface of the beads through electrostatic interactions 37 and molecular crowding. 38 Magnet fields are then applied to collect DNA-bound beads, effectively removing unwanted debris such as membrane lipids and proteins.…”

Shaking Device for Homogeneous Dispersion of Magnetic Beads in Droplet Microfluidics

“…Figure 5 b reports the results varying the volume of the sample (35 µL and 200 µL) and the bead concentration (4 × 10 8 beads/mL and 4 × 10 9 beads/mL). These values were chosen because they were typically used in many applications, such as immunoassays, mRNA and DNA purification, or extracellular vesicle isolation [ 17 , 46 , 47 ]. Figure 5 shows that, in all cases, activation of the shaking device promoted a good homogeneity of the bead distribution along the droplet train.…”

Draft genome sequence of multidrug-resistant Klebsiella pneumoniae Hakim-RU strain isolated from a patient with urinary tract infections in Bangladesh

“…The isolate was cultured in nutrient broth (HiMedia, India) at 37°C overnight, and its genomic DNA was extracted using the Qiagen DNA Mini Kit (QIAGEN, Hilden, Germany). Genomic DNA underwent enzymatic fragmentation using the NEBNext dsDNA Fragmentase Kit (NEB, MA, USA), followed by size selection with SPRI beads ( 15 ). A sequencing library was generated using the Nextera DNA Flex Library Prep Kit (Illumina, San Diego, CA, USA), and the library was sequenced with 2 × 150 paired-end reads on the Illumina NextSeq2000 platform.…”