Bulked oligo-FISH for chromosome painting and chromosome barcoding

Beránková D., Hřibová E.
Plant cytogenetics and cytogenomics: Methods and Protocols 2672: 445-463, 2023

Keywords: Oligo-based probe, mitotic metaphase chromosome, meiotic pachytene chromosome, oligo painting, fluorescence in situ hybridization
Abstract: 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 chromo-somes, 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, fluores-cently 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.).
DOI: doi.org/10.1007/978-1-0716-3226-0_27
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IEB authors: Denisa Beránková, Eva Hřibová