A comparison of DNA fragmentation methods − Applications for the biochip technology

Abstract

The efficiency of hybridization signal detection in a biochip is affected by the method used for test DNA preparation, such as fragmentation, amplification and fluorescent labelling. DNA fragmentation is the commonest methods used and it is recognised as a critical step in biochip analysis. Currently methods used for DNA fragmentation are based either on sonication or on the enzymatic digestion. In this study, we compared the effect of different types of enzymatic DNA fragmentations, using DNase I to generate ssDNA breaks, NEBNext dsDNA fragmentase and SaqAI restrictase, on DNA labelling. DNA from different Desulfovibrio species was used as a substrate for these enzymes. Of the methods used, DNA fragmented by NEBNext dsDNA Fragmentase digestion was subsequently labelled with the greatest efficiency. As a result of this, the use of this enzyme to fragment target DNA increases the sensitivity of biochip-based detection significantly, and this is an important consideration when determining the presence of targeted DNA in ecological and medical samples.

Recent Advances in Strategies for the Cloning of Natural Product Biosynthetic Gene Clusters

“…Regardless of the cloning method used, sufficient amounts of DNA fragments are indispensable. Therefore, the preparation of high-quality and high-molecular-weight DNA fragments is recognized as a critical step in gene cluster cloning ( Sapojnikova et al, 2017 ).…”

Properties and efficient scrap-and-build repairing of mechanically sheared 3’ DNA ends

“…The repair or polishing of DNA termini is a crucial step in the handling of DNA fragments, especially those generated by mechanical shearing, such as sonication, to efficiently conduct subsequent enzymatic reactions. DNA sonication 1 has been widely used for random subcloning of DNA for shotgun DNA sequence analysis 2 , the preparation of next-generation sequencing (NGS) libraries for genomic and meta-genomic sequencing 3–5 , Tn-seq analysis 6 , microarray analysis 7 , and ChIP analysis for mapping binding sites of DNA binding proteins 8 .…”