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利用Hi-C测序技术改进基因组组装及体外验证L.中差异表达基因

Exploitation of Hi-C sequencing for improvement of genome assembly and in-vitro validation of differentially expressing genes in L.

作者信息

Jalali Saakshi, Kancharla Nagesh, Yepuri Vijay, Arockiasamy Savarimuthu

机构信息

Agronomy Division, Reliance Technology Group, Reliance Industries Limited, Navi Mumbai, Maharashtra India.

出版信息

3 Biotech. 2020 Mar;10(3):91. doi: 10.1007/s13205-020-2082-0. Epub 2020 Feb 4.

Abstract

is one of the major sources of renewable energy due to potential use of its oil as a biofuel. The genome of this crop is constituted by the high content of repetitive elements. We employed the Hi-C proximity ligation technique to re-scaffold our existing hybrid genome assembly of an elite genotype (RJC1) developed using Illumina and Pacbio technologies. We assembled 99.81% of non-truncated reads to achieve 266.80 Mbp of the genome with an N50 value of 1.58 Mb. Furthermore, we compared the efficiency of Hi-C-augmented genome assembly with the hybrid genome assembly and observed a ~ 50% reduction in scaffolds and a tenfold increase in the N50 value. The gene ontology analysis revealed the identification of terms for molecular function (45.52%), cellular component (33.47%), and biological function (20.99%). Comparative genomic analysis of 13-plant species showed the conservation of 414 lipid metabolizing genes identified in the KEGG pathway analysis. Differential gene expression (DGE) studies were conducted in the healthy and Jatropha mosaic virus-infected leaves via RNA-seq analysis and observed gene expression changes for 2185 genes. Out of these, we observed 546 genes having more than two-fold change of transcript level and among these 259 genes were down-regulated and 287 genes were up-regulated. To validate RNA-seq data, two DEGs were selected for gene expression analysis using qRT-PCR and the data was in correlation with in silico results. RNA-seq analysis further shows the identification of some of the candidate genes and may be useful to develop JMV resistant plants after functional validation. This Hi-C genome assembly provides a detailed accurate reference genome which could be utilized to improve Jatropha and other economically important Euphorbiaceae family members.

摘要

由于其油有作为生物燃料的潜在用途,它是可再生能源的主要来源之一。这种作物的基因组由高含量的重复元件构成。我们采用Hi-C邻近连接技术对我们现有的使用Illumina和Pacbio技术开发的优良基因型(RJC1)的杂交基因组组装进行重新构建。我们组装了99.81%的非截断读段,获得了266.80 Mbp的基因组,N50值为1.58 Mb。此外,我们将Hi-C增强的基因组组装效率与杂交基因组组装进行了比较,发现支架数量减少了约50%,N50值增加了十倍。基因本体分析揭示了分子功能(45.52%)、细胞成分(33.47%)和生物学功能(20.99%)相关术语的识别。对13种植物的比较基因组分析表明,在KEGG通路分析中鉴定出的414个脂质代谢基因具有保守性。通过RNA-seq分析在健康和麻风树花叶病毒感染的叶片中进行了差异基因表达(DGE)研究,观察到2185个基因的表达变化。其中,我们观察到546个基因的转录水平变化超过两倍,其中259个基因下调,287个基因上调。为了验证RNA-seq数据,选择了两个差异表达基因进行qRT-PCR基因表达分析,数据与计算机模拟结果相关。RNA-seq分析进一步显示了一些候选基因的鉴定,在功能验证后可能有助于培育抗JMV的植物。这种Hi-C基因组组装提供了一个详细准确的参考基因组,可用于改良麻风树和其他经济上重要的大戟科植物。

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