Development of a fast and efficient ultrasonic-based strategy for DNA fragmentation

• Published in: Talanta (Oxford) 2010-05, Vol.81 (3), p.881-886
• Main Authors: Larguinho, Miguel, Santos, Hugo M., Doria, Gonçalo, Scholz, H., Baptista, Pedro V., Capelo, José L.
• Format: Article
• Language: English
• Subjects: Analytical chemistry; beta-Globins - genetics; Chemistry; DNA - analysis; DNA Fragmentation; DNA Restriction Enzymes - chemistry; Electrophoresis, Agar Gel - methods; Equipment Design; Exact sciences and technology; Humans; Ions; Oligonucleotides - chemistry; Plasmids - metabolism; Restriction enzyme; Sample preparation; Sonication; Sonoreactor; Sulfhydryl Compounds; Time Factors; Ultrasonics; Ultrasound
• ISSN: 0039-9140; 1873-3573
• DOI: 10.1016/j.talanta.2010.01.032

Several ultrasound-based platforms for DNA sample preparation were evaluated in terms of effective fragmentation of DNA (plasmid and genomic DNA)—ultrasonic probe, sonoreactor, ultrasonic bath and the newest Vialtweeter device. The sonoreactor showed the best efficiency of DNA fragmentation while simultaneously assuring no cross-contamination of samples, and was considered the best ultrasonic tool to achieve effective fragmentation of DNA at high-throughput and avoid sample overheating. Several operation variables were studied—ultrasonication time and amplitude, DNA concentration, sample volume and sample pre-treatment—that allowed optimisation of a sonoreactor-based strategy for effective DNA fragmentation. Optimal operating conditions to achieve DNA fragmentation were set to 100% ultrasonic amplitude, 100 μL sample volume, 8 min ultrasonic treatment (2 min/sample) for a DNA concentration of 100 μg mL −1. The proposed ultrasonication strategy can be easily implemented in any laboratory setup, providing fast, simple and reliable means for effective DNA sample preparation when fragmentation is critical for downstream molecular detection and diagnostics protocols.