Development of Oligo-GISH kits for efficient detection of chromosomal variants in peanut

Oligo probe staining is a low-cost and efficient chromosome identification technique. In this study, oligo genomic in situ hybridization (Oligo-GISH) technology was established in peanut. Peanut A and B subgenome-specific interspersed repeat (IR) oligo probe sets were developed based on clustering and electronic localization of tandem repeat sequences in the reference genome of Tifrunner. The Oligo-GISH kit was then used to perform staining of 15 Arachis species. The A-subgenome probe set stained the chromosomes of A- and E-genome Arachis species, the B-subgenome probe set stained those of B-, F-, K-, and E-genome species, and neither set stained those of H-genome species. These results indicate the relationships among the genomes of these Arachis species. The Oligo-GISH kit was also used for batch staining of the chromosomes of 389 seedlings from the irradiated M1 generation, allowing 67 translocation and deletion lines to be identified. Subsequent Oligo-FISH karyotyping, FISH using single-copy probe libraries, and trait investigation identified seven homozygous chromosomal variants from the M3 generation and suggested that there may be genes on chromosome 4B controlling seed number per pod. These findings demonstrate that the IR probe sets and method developed in this study can facilitate research on distant hybridization and genetic improvement in peanut.