Supplementary Material for: Characterization of Each St and Y Genome Chromosome of <b><i>Roegneria grandis</i></b> Based on Newly Developed FISH Markers

The genera of the tribe Triticeae (family Poaceae), constituting many economically important plants with abundant genetic resources, carry genomes such as St, H, P, and Y. The genome symbol of Roegneria C. Koch (Triticeae) is StY. The St and Y genomes are crucial in Triticeae, and tetraploid StY species participate extensively in polyploid speciation. Characterization of St and Y nonhomologous chromosomes in StY-genome species could help understand variation in the chromosome structure and differentiation of StY-containing species. However, the high genetic affinity between St and Y genome and the deficiency of a complete set of StY nonhomologous probes limit the identification of St and Y genomes and variation of chromosome structures among Roegneria species. We aimed to identify St- and Y-enhanced repeat clusters and to study whether homoeologous chromosomes between St and Y genomes could be accurately identified due to high affinity. We employed comparative genome analyses to identify St- and Y-enhanced repeat clusters and generated a FISH-based karyotype of R. grandis (Keng), one of the taxonomically controversial StY species, for the first time. We explored 4 novel repeat clusters (StY_34, StY_107, StY_90, and StY_93), which could specifically identify individual St and Y nonhomologous chromosomes. The clusters StY_107 and StY_90 could identify St and Y addition/substitution chromosomes against common wheat genetic backgrounds. The chromosomes V_St, VII_St, I_Y, V_Y, and VII_Y displayed similar probe distribution patterns in the proximal region, indicating that the high affinity between St and Y genome might result from chromosome rearrangements or transposable element insertion among V_St/Y, VII_St/Y, and I_Y chromosomes during allopolyploidization. Our results can be used to employ FISH further to uncover the precise karyotype based on colinearity of Triticeae species by using the wheat karyotype as reference, to analyze diverse populations of the same species to understand the intraspecific structural changes, and to generate the karyotype of different StY-containing species to understand the interspecific chromosome variation.禾本科（Poaceae）小麦族（Triticeae）各属包含众多具有丰富遗传资源的经济重要性植物，其基因组涵盖St、H、P、Y等类型。鹅观草属（Roegneria C. Koch，隶属于小麦族）的基因组符号为StY。St与Y基因组在小麦族中具有关键作用，四倍体StY基因组物种广泛参与多倍体物种形成过程。对StY基因组物种中St、Y非同源染色体的特征解析，有助于理解含StY基因组物种的染色体结构变异与分化情况。然而，St与Y基因组间极高的遗传同源性，以及完整的StY非同源染色体探针组的缺乏，限制了鹅观草属物种中St、Y基因组的识别以及染色体结构变异的解析。本研究旨在筛选St、Y基因组富集的重复序列簇，并探究在高遗传同源性背景下，能否精准识别St与Y基因组间的部分同源染色体。本研究通过比较基因组分析筛选得到St、Y基因组富集的重复序列簇，并首次构建了分类学上存在争议的StY基因组物种——大鹅观草（R. grandis (Keng)）的基于荧光原位杂交（FISH）的核型。本研究筛选得到4个新型重复序列簇（StY_34、StY_107、StY_90与StY_93），它们可特异性识别单条St或Y非同源染色体。其中StY_107与StY_90簇可在普通小麦遗传背景下识别St、Y基因组的附加/替换染色体。染色体V_St、VII_St、I_Y、V_Y与VII_Y的近着丝粒区域展现出相似的探针分布模式，这表明St与Y基因组间的高遗传同源性，可能源于异源多倍化过程中V_St/Y、VII_St/Y及I_Y染色体间发生的染色体重排或转座元件插入事件。本研究结果可为后续研究提供支撑：以小麦核型为参照，基于小麦族物种的共线性关系，通过荧光原位杂交技术解析精准核型；分析同一物种的不同种群以理解种内结构变异；以及构建不同含StY基因组物种的核型，以解析种间染色体变异。