Single copy oligonucleotide fluorescence in situ hybridization probe design platforms: Development, application and evaluation

Abstract

Oligonucleotides fluorescence in situ hybridization (Oligo‐FISH) is an emerging technology and is an important tool in research areas such as detection of chromosome variation, identification of allopolyploid, and deciphering of three‐dimensional (3D) genome structures. Based on the demand for highly efficient oligo probes for oligo‐FISH experiments, increasing numbers of tools have been developed for probe design in recent years. Obsolete oligonucleotide design tools have been adapted for oligo‐FISH probe design because of their similar considerations. With the development of DNA sequencing and large‐scale synthesis, novel tools have been designed to increase the specificity of designed oligo probes and enable genome‐scale oligo probe design, which has greatly improved the application of single copy oligo‐FISH. Despite this, few studies have introduced the development of the oligo‐FISH probe design tools and their application in FISH experiments systematically. Besides, a comprehensive comparison and evaluation is lacking for the available tools. In this review, we provide an overview of the oligo‐FISH probe design process, summarize the development and application of the available tools, evaluate several state‐of‐art tools, and eventually provide guidance for single copy oligo‐FISH probe design.