Rapid Nucleic Acid Extraction and Purification Using a Miniature Ultrasonic Technique

Abstract

Miniature ultrasonic lysis for biological sample preparation is a promising technique for efficient and rapid extraction of nucleic acids and proteins from a wide variety of biological sources. Acoustic methods achieve rapid, unbiased, and efficacious disruption of cellular membranes while avoiding the use of harsh chemicals and enzymes, which interfere with detection assays. In this work, a miniature acoustic nucleic acid extraction system is presented. Using a miniature bulk acoustic wave (BAW) transducer array based on 36° Y-cut lithium niobate, acoustic waves were coupled into disposable laminate-based microfluidic cartridges. To verify the lysing effectiveness, the amount of liberated ATP and the cell viability were measured and compared to untreated samples. The relationship between input power, energy dose, flow-rate, and lysing efficiency were determined. DNA was purified on-chip using three approaches implemented in the cartridges: a silica-based sol-gel silica-bead filled microchannel, nucleic acid binding magnetic beads, and Nafion-coated electrodes. Using E. coli, the lysing dose defined as ATP released per joule was 2.2× greater, releasing 6.1× more ATP for the miniature BAW array compared to a bench-top acoustic lysis system. An electric field-based nucleic acid purification approach using Nafion films yielded an extraction efficiency of 69.2% in 10 min for 50 µL samples.

Active Microbiome Structure and Functional Analyses of Freshwater Benthic Biofilm Samples Influenced by RNA Extraction Methods

“…Following a cleanup step comprising the truncation of barcode and primer sequences, the paired-end reads were merged using FLASH (Magoc and Salzberg, 2011) to obtain raw reads that were later subjected to quality filtering to obtain high-quality clean reads (Bokulich et al, 2013) according to the QIIME quality control process (Caporaso et al, 2010b). To obtain effective tags, the UCHIME algorithm was used to compare the cleaned read sequences with a reference Gold database (Edgar et al, 2011) and thus enable the removal of chimera sequences.…”

“…Quantitative metabolic mapping of the metatranscriptome profiles was performed by mapping the KEGG annotations of identified protein sequences into metabolic pathway maps, using the iPath v2 module (Bokulich et al, 2013). In these maps, the gene expression associated with a metabolic pathway was indicated by the line width, which was determined using the log2 read count values of KEGG-annotated proteins.…”

Advances in point-of-care nucleic acid extraction technologies for rapid diagnosis of human and plant diseases

“…Several on-chip cell lysis techniques such as chemical lysis ( Ma et al, 2019 ; Yoon et al, 2018 ), mechanical lysis (J. Choi et al, 2015 ; Mahalanabis et al, 2009 ), electrical lysis ( Hügle et al, 2018 ; Kim et al, 2009 ; Nan et al, 2014 ), ultrasonic lysis ( Branch et al, 2017 ), thermal lysis ( Leiske et al, 2015 ; Wang et al, 2011 ; Zhang et al, 2016 ), and enzymatic lysis ( Lounsbury et al, 2013 ; Petralia et al, 2013 ) have been demonstrated for rapid lysis of human and pathogen cells.…”

“…For POC applications, solid-phase extraction (SPE) is more widely used for nucleic acid isolation and purification ( Kim et al, 2010 ; Price et al, 2009 ; Reinholt and Baeumner, 2014 ). In SPE, cell lysate is mixed with or passed through solid-phase materials, such as filter papers (R. Tang et al, 2017a ), silica membranes or beads ( Branch et al, 2017 ), polymer resins ( Byrnes et al, 2013 ), organic ligands ( Jin et al, 2017 ), nanomaterials (H. Liu et al, 2018 ), or magnetic particles ( Fu et al, 2018 ; Kang et al, 2017 ; Neto et al, 2017 ) to selectively bind nucleic acids at a pH lower than 7.5 ( Yoon et al, 2018 ), remove impurities, and elute DNA or RNA molecules from the solid phase at a higher pH (~pH 8).…”

Application of Microfluidic Methodology for the Analysis of DNA

“…Following sample addition, cell lysis is commonly required to release nucleic acids, however, chemical or enzymatic cell lysis methods can have adverse effects on downstream processes in integrated systems. Branch et al [ 5 ] have evaluated a miniature bulk acoustic wave transducer array to overcome these limitations, and demonstrated extraction efficiencies of nearly 70% in just 10 min.…”