利用基因體學策略探勘甘藍耐淹水相關基因

Abstract

甘藍 (Brassica oleracea L. var. capitata) 為臺灣重要蔬菜作物，農業生產上容易遭受降雨造成的淹水逆境影響，造成嚴重經濟損失。因此，了解甘藍耐淹基礎分子機制對於品種育成具重要應用價值。本研究欲透過不同次世代定序技術，針對本研究室先前建立之8個與甘藍耐淹相關數量性狀基因座區間內，分析耐淹品種甘藍‘大蕊’ (‘Fuyudori’) 與不耐淹品種甘藍‘228’之間的基因體序列及轉錄體差異，探討可能導致兩品種耐淹能力不同之關聯性。以目標區域定序策略針對甘藍株高相關數量性狀基因座C7_44554117總長500 kb之區間範圍進行定序，共獲得20個預測為高影響變異之基因，其中，‘大蕊’與‘228’中分別具有6個與10個變異存在於單一品種之基因。全基因體定序則對‘大蕊’與‘228’兩甘藍品種進行分析，於8個甘藍耐淹數量性狀基因座區間內獲得‘大蕊’中的299個與‘228’中的244個預測為高影響變異之基因，其中，‘大蕊’與‘228’中分別具有96與41個變異存在於單一品種之基因。根據變異資訊與基因功能註解，選出29個與耐淹性相關之基因。另外利用本研究室既有之轉錄體定序數據，分析甘藍於淹水期間之基因表現變化。自8個甘藍耐淹數量性狀基因座區間範圍內，獲得90個於淹水處理下與兩品種間具有差異表現之基因，並選出26個與耐淹性相關之基因。整合三策略之資訊後獲得12個具品種特異性變異與差異表現之基因，其中抗氧化酵素相關基因PER50在兩甘藍品種淹水期間被誘導表現，且‘大蕊’表現提升較‘228’多，推測‘大蕊’具有較佳於淹水逆境下維持ROS平衡的能力；此外，負責氧化細胞分裂素之基因CKX7，在‘大蕊’中具有位於蛋白質結構域內的序列變異，可能因此改變其蛋白質功能進而影響其淹水耐受性。整體而言，本研究提供甘藍耐淹調控機制解析之資訊，期望助於提升耐淹甘藍品種之育成效率。

Cabbage (Brassica oleracea L. var. capitata) is one of the most important vegetable crops in Taiwan. Flooding stress caused by heavy rainfall often affected its agricultural production, resulting in significant economic losses. Therefore, elucidating the molecular mechanisms underlying flooding tolerance in cabbage is crucial for breeding flood-tolerant cultivars. This study aims to investigate the genomic and transcriptomic differences within the flanking region of eight previously identified quantitative trait loci (QTLs) associated with flooding tolerance between a flooding-tolerant cabbage cultivar, ‘Fuyudori’, and a flooding-intolerant cultivar, ‘228’, based on next-generation sequencing (NGS) strategies to explore the potential molecular basis underlying their differential flooding tolerance. Targeted sequencing for a 500 kb flanking region of the QTL C7_44554117 that is associated with plant height trait identified a total of 20 genes with predicted high-impact variants, among which 6 and 10 genes were found exclusively in ‘Fuyudori’ and ‘228’, respectively. Whole-genome sequencing (WGS) analysis within the flanking region of eight QTL identified 299 and 244 genes with predicted high-impact variants in ‘Fuyudori’ and ‘228’, respectively, including 96 and 41 genes with unique variants. Based on variant information and gene function annotation, 29 candidate genes associated with cabbage flooding tolerance were selected. Additionally, 90 differentially expressed genes (DEGs) under flooding treatment within the same QTL regions were identified based on the existing RNA sequencing (RNA-seq) in our laboratory, of which 26 were considered associated with flooding tolerance. By integrating the results of the three strategies, 12 candidate genes were identified as harboring both cultivar-specific variants and differential expression under flooding stress. Among these, the antioxidant-related gene PER50 was induced during flooding in both cultivars, with a higher expression increase observed in ‘Fuyudori’, suggesting its involvement in maintaining reactive oxygen species (ROS) homeostasis under stress conditions. Furthermore, the CKX7 gene, which encodes a cytokinin dehydrogenase, harbored a sequence variant within a functional domain in ‘Fuyudori’, potentially affecting its protein function and consequently influencing flooding tolerance. In conclusion, this study provides valuable insights into the molecular regulation mechanisms of flooding tolerance in cabbage. It identifies potential candidate genes for future functional validation and molecular marker-assisted selection, thereby contributing to the development of flood-tolerant cabbage cultivars.