基于BSA-seq的西瓜白色果皮基因定位分析

Gene mapping analysis of white fruit rind trait in watermelon based on BSA-seq

摘要

以白色果皮西瓜品种三白瓜和墨绿色果皮材料LN1-1为试验材料，构建遗传群体，系统解析西瓜白色果皮遗传机制，定位调控该性状的关键基因，为白色果皮西瓜分子标记辅助育种提供理论支撑。以三白瓜为母本、LN1-1为父本构建四世代遗传群体，调查F₂代群体果皮颜色表型并进行遗传分析；利用BSA-seq（分离群体混合池测序）技术对F₂代白色与非白色果皮极端个体混池进行测序，结合生物信息学分析定位关键候选基因，通过序列比对、系统进化树分析验证基因功能关联性。结果表明，F₂代225单株中，白色果皮60株、非白色果皮165株，分离比符合理论比1:3（χ²=0.33，P＞0.05），表明白色果皮受单隐性核基因控制。BSA-seq测序分析将该基因定位在9号染色体11 793 001~14 532 000 bp（2.74 Mb）区间内。该区间78个基因中，Cla97C09G175170（与APRR2同源）被预测为关键候选基因。序列分析显示，该基因编码区无变异，仅在第9内含子存在1个与白/非白表型共分离的G→A关键SNP位点；系统进化树分析证实其编码蛋白与冬瓜、黄瓜、番茄等作物的同源蛋白序列高度保守。结合APRR2基因的保守功能，推测该内含子突变可能通过影响基因表达进而阻碍叶绿体发育并导致叶绿素完全缺失，从而呈现白色果皮。本研究成功定位并预测Cla97C09G175170（与APRR2同源）为控制西瓜白色果皮性状的关键候选基因，为西瓜白色果皮性状的分子标记辅助育种及基因功能验证奠定了重要理论基础。

Abstract

This study used the watermelon with white peel variety Sanbai Gua and the dark green peel material LN1-1 as parents to construct a genetic population, systematically dissect the genetic mechanism of white peel in watermelon, map the key genes regulating this trait, with the aim of providing a theoretical basis for molecular marker-assisted breeding of watermelon with white peel. Using Sanbai Gua as the female parent and LN1-1 as the male parent, a four-generation population was constructed. The peel color phenotype of the F₂ population was investigated, and subjected to genetic analysis. Bulked segregant analysis combined with whole-genome resequencing (BSA-seq) technology was employed to sequence the extreme pools of white and non-white peel individuals in the F₂ generation, and bioinformatics analysis was used to map key candidate genes, followed by sequence alignment and phylogenetic tree analysis to verify gene functional associations. The results showed that among the 225 F₂ plants, 60 displayed white peel and 165 showed non-white peel, with a segregation ratio fitting the theoretical 1∶3 ratio (χ²=0.33, P＞0.05), indicating that white peel trait is controlled by a single recessive nuclear gene. BSA-seq localized the gene to a 2.74 Mb interval on chromosome 9 (11 793 001-14 532 000 bp), containing 78 genes, among whichCla97C09G175170 (a homolog of APRR2) was predicted as the key candidate gene. Sequence analysis revealed no variation in the coding region of this gene, with only one critical G→A SNP within the 9th intron that co-segregated with the white/non-white phenotype. Phylogenetic tree analysis showed that its encoded protein is highly conserved with homologous protein sequences from other crops such as wax gourd, cucumber, and tomato. Combined with the known function of theAPRR2 gene, it is speculated that this intronic mutation may affect gene expression, thereby impairing chloroplast development and leading to complete loss of chlorophyll, ultimately resulting in the white peel phenotype. This study successfully mapped and predicted Cla97C09G175170 as the key candidate gene controlling watermelon white peel, laying an important theoretical foundation for further gene functional validation, and molecular marker-assisted breeding.

关键词

西瓜 / 白色果皮 / 遗传分析 / BSA-seq / APRR2基因

Key words

Watermelon / White peel / Genetic analysis / BSA-seq / APRR2 gene

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基金资助

辽宁省教育厅基本科研项目(LJKQZ20222426)