Poinsettia is an economically important ornamental potted plant in which certain bract colour variants are often obtained by mutation breeding. Previously, in poinsettia, we identified Bract1, a GST gene involved in the sequestration and transport of anthocyanins to the vacuole. This gene carries a short, highly mutable 4-bp repeat in its coding region. Loss of one repeat unit leads to a loss of function for Bract1, and in homozygous mutants, anthocyanin-based coloration is absent, resulting in white or cream-coloured bracts. Although mutation induction through ionizing radiation leads to a high frequency of mutations in Bract1, mutants are difficult to obtain from homozygous red genotypes. In this study, we used Bract1-specific amplicon sequencing as a tool to identify mutations in pools of tissues, which enabled the detection of mutations in dilutions of up to one mutant in 50 nonmutated samples. This approach enabled efficient screening of recalcitrant homozygous genotypes for mutated alleles and the reduction of the mutation load in the application of ionizing radiation in mutation breeding programmes.

中文翻译：

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开发用于检测一品红 (Euphorbia pulcherrima) 育种计划中低频突变的多重扩增子测序分析

一品红是一种经济上重要的观赏盆栽植物，其中某些苞片颜色变种通常通过突变育种获得。以前，我们在一品红中鉴定了Bract1，这是一种GST基因，参与了花青素向液泡的隔离和转运。该基因在其编码区携带一个短的、高度可变的 4 bp 重复序列。一个重复单元的丢失导致Bract1的功能丧失，并且在纯合突变体中，不存在基于花青素的着色，导致白色或奶油色的苞片。尽管通过电离辐射诱导突变导致Bract1突变频率很高，但很难从纯合红色基因型中获得突变体。在本研究中，我们使用Bract1-特定的扩增子测序作为一种工具来识别组织库中的突变，这使得在 50 个非突变样本中最多一个突变体的稀释液中检测突变成为可能。这种方法能够有效筛选突变等位基因的顽固纯合基因型，并减少在突变育种计划中应用电离辐射时的突变负荷。