Recently developed bulked oligo-FISH is a highly versatile method, which is applicable in any plant species with an assembled genome sequence. This technique allows in situ identification of individual chromosomes, large chromosomal rearrangements, comparative karyotype analysis, or even the reconstruction of the three-dimensional organization of the genome. The method is based on the identification of thousands of short oligonucleotides, unique to specific genome regions, which are synthesized in parallel, fluorescently labeled and used as probes for FISH. In this chapter, we propose a detailed protocol for amplification and labeling of single-stranded oligo-based painting probes from so-called MYtags immortal libraries, the preparation of mitotic metaphase and meiotic pachytene chromosome spreads, and a protocol for the fluorescence in situ hybridization procedure using the synthetic oligo probes. The proposed protocols are demonstrated for banana (Musa spp.).

Institute of Experimental Botany of the Czech Academy of Sciences Centre of Plant Structural and Functional Genomics Olomouc Czech Republic

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https://github.com/forrestzhang/Chorus

Protocol for preparing single-stranded labeled probes from a MYtags immortal library (Daicel Arbor Biosciences, Ann Arbor, Michigan, USA, 2016)

Labeling protocol for MYtags immortal libraries (Daicel Arbor Biosciences, Ann Arbor, Michigan, USA, 2018, version 1.5)

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Striking variation in chromosome structure within Musa acuminata subspecies, diploid cultivars, and F1 diploid hybrids