Nagano Vegetable and Ornamental Crops Experiment Station, Tokoo 1066-1, Souga, Shiojiri, Nagano, 399-6461, Japan.
Department of Bioresource Development, Tokyo University of Agriculture, Funako 1737, Atsugi, Kanagawa, 243-0034, Japan.
Plant Cell Rep. 2024 Jan 11;43(2):35. doi: 10.1007/s00299-023-03124-4.
We identified LsMybW as the allele responsible for the shift in color from black to white seeds in wild ancestors of lettuce to modern cultivars. Successfully selected white seeds are a key agronomic trait for lettuce cultivation and breeding; however, the mechanism underlying the shift from black-in its wild ancestor-to white seeds remains uncertain. We aimed to identify the gene/s responsible for white seed trait in lettuce. White seeds accumulated less proanthocyanidins than black seeds, similar to the phenotype observed in Arabidopsis TT2 mutants. Genetic mapping of a candidate gene was performed with double-digest RAD sequencing using an F population derived from a cross between "ShinanoPower" (white) and "Escort" (black). The white seed trait was controlled by a single recessive locus (48.055-50.197 Mbp) in linkage group 7. Using five PCR-based markers and numerous cultivars, eight candidate genes were mapped in the locus. Only the LG7_v8_49.251Mbp_HinfI marker, employing a single-nucleotide mutation in the stop codon of Lsat_1_v5_gn_7_35020.1, was completely linked to seed color phenotype. In addition, the coding region sequences for other candidate genes were identical in the resequence analysis of "ShinanoPower" and "Escort." Therefore, we proposed Lsat_1_v5_gn_7_35020.1 as the candidate gene and designated it as LsMybW (Lactuca sativa Myb White seeds), an ortholog encoding the R2R3-MYB transcription factor in Arabidopsis. When we validated the role of LsMybW through genome editing, LsMybW knockout mutants harboring an early termination codon showed a change in seed color from black to white. Therefore, LsMybW was the allele responsible for the shift in seed color. The development of a robust marker for marker-assisted selection and identification of the gene responsible for white seeds have implications for future breeding technology and physiological analysis.
我们鉴定出 LsMybW 是导致生菜野生祖先的黑色种子变为白色种子的等位基因。成功选育的白色种子是生菜种植和培育的关键农艺性状；然而，其野生祖先的黑色种子变为白色种子的机制尚不清楚。我们旨在鉴定生菜中白色种子性状的相关基因。白色种子积累的原花青素少于黑色种子，与拟南芥 TT2 突变体观察到的表型相似。利用来自“ShinanoPower”（白色）和“Escort”（黑色）杂交的 F 群体，通过双酶切 RAD 测序对候选基因进行遗传作图。白色种子性状由第 7 连锁群上的单个隐性位点（48.055-50.197 Mb）控制。利用 5 个基于 PCR 的标记和多个品种，将 8 个候选基因定位在该位点。只有 LG7_v8_49.251Mbp_HinfI 标记，其在 Lsat_1_v5_gn_7_35020.1 的终止密码子处发生单核苷酸突变，与种子颜色表型完全连锁。此外，在“ShinanoPower”和“Escort”的重测序分析中，其他候选基因的编码区序列完全相同。因此，我们提出 Lsat_1_v5_gn_7_35020.1 是候选基因，并将其命名为 LsMybW（生菜 R2R3-MYB 转录因子 White seeds），它是拟南芥 R2R3-MYB 转录因子的同源基因。当我们通过基因组编辑验证 LsMybW 的作用时，携带提前终止密码子的 LsMybW 敲除突变体的种子颜色从黑色变为白色。因此，LsMybW 是导致种子颜色改变的等位基因。该研究为标记辅助选择和鉴定白色种子相关基因开发了一种强大的标记，对未来的育种技术和生理分析具有重要意义。
