National Agriculture and Food Research Organization Institute of Fruit and Tea Tree Science, Shimizu, Shizuoka, Japan.
Laboratory of Biometry and Bioinformatics, Department of Agricultural and Environmental Biology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo, Tokyo, Japan.
PLoS One. 2021 Feb 4;16(2):e0246468. doi: 10.1371/journal.pone.0246468. eCollection 2021.
To enrich carotenoids, especially β-cryptoxanthin, in juice sac tissues of fruits via molecular breeding in citrus, allele mining was utilized to dissect allelic variation of carotenoid metabolic genes and identify an optimum allele on the target loci characterized by expression quantitative trait (eQTL) analysis. SNPs of target carotenoid metabolic genes in 13 founders of the Japanese citrus breeding population were explored using the SureSelect target enrichment method. An independent allele was determined based on the presence or absence of reliable SNPs, using trio analysis to confirm inheritability between parent and offspring. Among the 13 founders, there were 7 PSY alleles, 7 HYb alleles, 11 ZEP alleles, 5 NCED alleles, and 4 alleles for the eQTL that control the transcription levels of PDS and ZDS among the ancestral species, indicating that some founders acquired those alleles from them. The carotenoid composition data of 263 breeding pedigrees in juice sac tissues revealed that the phenotypic variance of carotenoid composition was similar to that in the 13 founders, whereas the mean of total carotenoid content increased. This increase in total carotenoid content correlated with the increase in either or both β-cryptoxanthin and violaxanthin in juice sac tissues. Bayesian statistical analysis between allelic composition of target genes and carotenoid composition in 263 breeding pedigrees indicated that PSY-a and ZEP-e alleles at PSY and ZEP loci had strong positive effects on increasing the total carotenoid content, including β-cryptoxanthin and violaxanthin, in juice sac tissues. Moreover, the pyramiding of these alleles also increased the β-cryptoxanthin content. Interestingly, the offset interaction between the alleles with increasing and decreasing effects on carotenoid content and the epistatic interaction among carotenoid metabolic genes were observed and these interactions complexed carotenoid profiles in breeding population. These results revealed that allele composition would highly influence the carotenoid composition in citrus fruits. The allelic genotype information for the examined carotenoid metabolic genes in major citrus varieties and the trio-tagged SNPs to discriminate the optimum alleles (PSY-a and ZEP-e) from the rest would promise citrus breeders carotenoid enrichment in fruit via molecular breeding.
为了通过分子育种在柑橘果实的汁囊组织中富集类胡萝卜素，特别是β-隐黄质，利用等位基因挖掘技术对类胡萝卜素代谢基因的等位基因变异进行了剖析，并通过表达数量性状（eQTL）分析鉴定了目标基因座上的最优等位基因。使用 SureSelect 目标富集方法，对日本柑橘育种群体的 13 个供体的目标类胡萝卜素代谢基因的 SNP 进行了探索。基于可靠 SNP 的存在或不存在，使用三亲分析确定了一个独立的等位基因，以确认亲本与后代之间的可遗传性。在 13 个供体中，PSY 有 7 个等位基因，HYb 有 7 个等位基因，ZEP 有 11 个等位基因，NCED 有 5 个等位基因，控制前体物种 PDS 和 ZDS 转录水平的 eQTL 有 4 个等位基因，表明其中一些供体从它们那里获得了这些等位基因。在汁囊组织中，263 个育种谱系的类胡萝卜素组成数据表明，类胡萝卜素组成的表型方差与 13 个供体相似，而总类胡萝卜素含量的平均值增加。这种总类胡萝卜素含量的增加与汁囊组织中β-隐黄质和紫黄质的增加有关。贝叶斯统计分析表明，在 263 个育种谱系的目标基因的等位基因组成与类胡萝卜素组成之间，PSY 和 ZEP 基因座上的 PSY-a 和 ZEP-e 等位基因对增加汁囊组织中的总类胡萝卜素含量（包括β-隐黄质和紫黄质）有很强的正向作用。此外，这些等位基因的聚合也增加了β-隐黄质的含量。有趣的是，观察到了增加和减少类胡萝卜素含量的等位基因之间的抵消互作以及类胡萝卜素代谢基因之间的上位性互作，并使这些互作在育种群体中复杂化了类胡萝卜素图谱。这些结果表明，等位基因组成会高度影响柑橘果实中的类胡萝卜素组成。主要柑橘品种中所检查的类胡萝卜素代谢基因的等位基因基因型信息以及区分最优等位基因（PSY-a 和 ZEP-e）的三亲标记 SNP，将使柑橘育种家能够通过分子育种在果实中实现类胡萝卜素的富集。
