Integrated Transcriptome and Metabolome Identified Novel Candidate Genes Regulating Anthocyanin Biosynthesis in Colored-Grain Wheat (Triticum aestivum L.). Integrated Transcriptome and Metabolome Identified Novel Candidate Genes Regulating Anthocyanin Biosynthesis in Colored-Grain Wheat (Triticum aestivum L.)

Purpose: The present study aimed to investigate the anthocyanin components and identify relevant regulatory genes in purple wheat grain by carrying out transcriptome analyses. Methods: The seeds of purple grain wheat and white grain wheat were collected 30 days after flowering, and three biological replicates were set. Total RNA was isolated and purified using TRIzol reagent (Invitrogen, Carlsbad, CA, USA) following the manufacturer's procedure. The RNA amount and purity of each sample was quantified using NanoDrop ND-1000. Then synthesizing the fragmented RNA into cDNA through the action of reverse transcriptase, and finally obtaining acDNA library. At last, we performed the 2×150bp paired-end sequencing (PE150) on an Illumina Novaseq™ 6000 following the vendor's recommended protocol. Results: A total of 10440 diferentially expressed genes were signifcantly enriched by RNA sequencing. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed signifcantly enriched flavonoid biosynthesis and anthocyanin biosynthesis in CW_S versus W_S. And the ANS and UFGT genes were predicted as core genes in anthocyanin biosynthesis. Conclusions: Our study represents the detailed analysis of wheat grain transcriptomes, with biologic replicates, generated by RNA-seq technology. Through this study, we speculated that ANS and UFGT genes are the core genes of anthocyanin biosynthesis.The significant differences of these genes affect the synthesis of anthocyanins in wheat grains, and thus affect the grain color of wheat. Overall design: The mRNA profiles of white grain wheat (W_S) and purple grain wheat (CW_S) grains 30 days after flowering.

研究目的：本研究旨在通过转录组分析，探究紫粒小麦籽粒中的花青素成分，并鉴定相关调控基因。
试验方法：本研究采集开花后30天的紫粒小麦与白粒小麦种子，并设置3次生物学重复。按照厂商操作流程，使用TRIzol试剂（美国加利福尼亚州卡尔斯巴德Invitrogen公司）分离并纯化总RNA。采用NanoDrop ND-1000对每个样品的RNA总量及纯度进行定量检测。随后通过逆转录酶将片段化RNA反转录为cDNA，最终构建cDNA文库。最后，依照厂商推荐的实验方案，在Illumina NovaSeq™ 6000测序平台上完成2×150bp双端测序（PE150）。
试验结果：经RNA测序，共筛选得到10440个显著差异表达基因。基因本体（Gene Ontology, GO）与京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG）富集分析结果显示，在CW_S与W_S的比较组中，类黄酮生物合成及花青素生物合成通路显著富集。预测ANS与UFGT基因为花青素生物合成的核心调控基因。
研究结论：本研究借助RNA测序技术，结合生物学重复实验，完成了小麦籽粒转录组的详细解析。通过本研究推测，ANS与UFGT基因为花青素生物合成的核心基因；这些基因的表达差异会影响小麦籽粒中花青素的合成，进而改变小麦籽粒的颜色。
试验设计：采集开花后30天的白粒小麦（W_S）与紫粒小麦（CW_S）籽粒的mRNA表达谱。