TY - JOUR

T1 - An inexpensive, simple and effective method of genome DNA fragmentation for NGS libraries

AU - Kechin, Andrey

AU - Boldyreva, Darya

AU - Borobova, Viktoriya

AU - Boyarskikh, Ulyana

AU - Scherbak, Sergey

AU - Apalko, Svetlana

AU - Makarova, Maria

AU - Mosyakin, Nikolay

AU - Kaftyreva, Lidia

AU - Filipenko, Maxim

N1 - Publisher Copyright: © 2021 The Author(s) 2021. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

PY - 2021/11/1

Y1 - 2021/11/1

N2 - Next-generation sequencing (NGS)-library preparation for whole-genome sequencing (WGS) starts with DNA fragmentation, and sonication is a physical approach used most often due to its simplicity and reproducibility. However, the commercially available Covaris instrument has a high price for both the device and consumables. Here, we describe our in-house method of DNA shearing by sonication with small (100-600 µm) glass beads and an ultrasonic bath. The fragmentation conditions were optimized for the bacterial WGS with ∼550-bp fragment size (the ultrasonic bath water temperature 5-10°C, glass beads 0.06 g, the fragmentation time 50 s) and for human DNA with ∼250 bp (fragmentation with the same parameters for 4 min). Fragmentation results were compared with the Covaris instrument for preparing several bacterial NGS libraries for Illumina NGS platforms by several characteristics. We obtained close mean fragment lengths (523-623 versus 480-646), similar mono- and dinucleotide specificity of shearing, and comparable indicators of read alignment and de novo assembly for both methods. Thus, the described method is a new fast, and effective DNA fragmentation approach that can be used in different WGS applications.

AB - Next-generation sequencing (NGS)-library preparation for whole-genome sequencing (WGS) starts with DNA fragmentation, and sonication is a physical approach used most often due to its simplicity and reproducibility. However, the commercially available Covaris instrument has a high price for both the device and consumables. Here, we describe our in-house method of DNA shearing by sonication with small (100-600 µm) glass beads and an ultrasonic bath. The fragmentation conditions were optimized for the bacterial WGS with ∼550-bp fragment size (the ultrasonic bath water temperature 5-10°C, glass beads 0.06 g, the fragmentation time 50 s) and for human DNA with ∼250 bp (fragmentation with the same parameters for 4 min). Fragmentation results were compared with the Covaris instrument for preparing several bacterial NGS libraries for Illumina NGS platforms by several characteristics. We obtained close mean fragment lengths (523-623 versus 480-646), similar mono- and dinucleotide specificity of shearing, and comparable indicators of read alignment and de novo assembly for both methods. Thus, the described method is a new fast, and effective DNA fragmentation approach that can be used in different WGS applications.

KW - DNA fragmentation

KW - glass beads

KW - shearing

KW - sonication

KW - whole-genome sequencing

UR - http://www.scopus.com/inward/record.url?scp=85123226424&partnerID=8YFLogxK

UR - https://www.elibrary.ru/item.asp?id=47558877

U2 - 10.1093/jb/mvab089

DO - 10.1093/jb/mvab089

M3 - Article

C2 - 34382083

AN - SCOPUS:85123226424

VL - 170

SP - 675

EP - 681

JO - Journal of biochemistry

JF - Journal of biochemistry

SN - 0021-924X

IS - 5

ER -