Grupo de Investigación en Genética Aplicada (GIGA), Facultad de Ciencias Exactas Químicas y Naturales, Instituto de Biología Subtropical (IBS UNaM-CONICET), Universidad Nacional de Misiones, Jujuy 1745, CP3300, Posadas, Misiones, Argentina.
The Genome Analysis Centre, Norwich Research Park, Norwich, NR4 7UH, UK.
BMC Genomics. 2018 Dec 7;19(1):891. doi: 10.1186/s12864-018-5240-6.
The most common infusion in southern Latin-American countries is prepared with dried leaves of Ilex paraguariensis A. St.-Hil., an aboriginal ancestral beverage known for its high polyphenols concentration currently consumed in > 90% of homes in Argentina, in Paraguay and Uruguay. The economy of entire provinces heavily relies on the production, collection and manufacture of Ilex paraguariensis, the fifth plant species with highest antioxidant activity. Polyphenols are associated to relevant health benefits including strong antioxidant properties. Despite its regional relevance and potential biotechnological applications, little is known about functional genomics and genetics underlying phenotypic variation of relevant traits. By generating tissue specific transcriptomic profiles, we aimed to comprehensively annotate genes in the Ilex paraguariensis phenylpropanoid pathway and to evaluate differential expression profiles.
In this study we generated a reliable transcriptome assembly based on a collection of 15 RNA-Seq libraries from different tissues of Ilex paraguariensis. A total of 554 million RNA-Seq reads were assembled into 193,897 transcripts, where 24,612 annotated full-length transcripts had complete ORF. We assessed the transcriptome assembly quality, completeness and accuracy using BUSCO and TransRate; consistency was also evaluated by experimentally validating 11 predicted genes by PCR and sequencing. Functional annotation against KEGG Pathway database identified 1395 unigenes involved in biosynthesis of secondary metabolites, 531 annotated transcripts corresponded to the phenylpropanoid pathway. The top 30 differentially expressed genes among tissue revealed genes involved in photosynthesis and stress response. These significant differences were then validated by qRT-PCR.
Our study is the first to provide data from whole genome gene expression profiles in different Ilex paraguariensis tissues, experimentally validating in-silico predicted genes key to the phenylpropanoid (antioxidant) pathway. Our results provide essential genomic data of potential use in breeding programs for polyphenol content. Further studies are necessary to assess if the observed expression variation in the phenylpropanoid pathway annotated genes is related to variations in leaves' polyphenol content at the population scale. These results set the current reference for Ilex paraguariensis genomic studies and provide a substantial contribution to research and biotechnological applications of phenylpropanoid secondary metabolites.
在南美南部国家，最常见的输液是用巴拉圭冬青的干叶制成的，这是一种土著祖先饮料，以其高多酚浓度而闻名，目前在阿根廷、巴拉圭和乌拉圭 90%以上的家庭中都有饮用。整个省份的经济严重依赖于巴拉圭冬青的生产、收集和制造，这是第五种具有最高抗氧化活性的植物物种。多酚与相关的健康益处有关，包括强大的抗氧化特性。尽管它具有地区相关性和潜在的生物技术应用，但对表型变异相关特征的功能基因组学和遗传学知之甚少。通过生成组织特异性转录组谱，我们旨在全面注释 Ilex paraguariensis 苯丙素途径中的基因，并评估差异表达谱。
在这项研究中，我们基于 Ilex paraguariensis 不同组织的 15 个 RNA-Seq 文库，生成了一个可靠的转录组组装。总共 5.54 亿个 RNA-Seq reads 被组装成 193897 个转录本，其中 24612 个注释的全长转录本具有完整的 ORF。我们使用 BUSCO 和 TransRate 评估转录组组装的质量、完整性和准确性；通过 PCR 和测序实验验证 11 个预测基因来评估一致性。KEGG 途径数据库的功能注释鉴定出 1395 个与次生代谢物生物合成相关的 unigenes，531 个注释的转录本对应于苯丙素途径。组织中差异表达基因的前 30 个基因涉及光合作用和应激反应。然后通过 qRT-PCR 验证这些显著差异。
我们的研究首次提供了不同 Ilex paraguariensis 组织的全基因组基因表达谱数据，实验验证了苯丙素（抗氧化）途径中关键的基于计算机预测的基因。我们的结果提供了多酚含量育种计划潜在有用的重要基因组数据。进一步的研究有必要评估在苯丙素途径注释基因中观察到的表达变化是否与群体尺度上叶片多酚含量的变化有关。这些结果为 Ilex paraguariensis 基因组研究设定了当前的参考标准，并为苯丙素类次生代谢物的研究和生物技术应用提供了重要贡献。
