Design and validation of microarray for chickpea (Cicer arietinum) based on the analysis of transcriptome data [gene expression]. Cicer arietinum

Microarrays have increasingly become a powerful tool for high throughput gene-expression studies and discovery of novel biomarker genes. Developed for a large number of organisms, including plants, microarrays are commonly performed for species that have sequenced data, for performing gene expression analysis, miRNA profiling, comparative genomic hybridization (CGH), ChIP-on-chip and SNP analysis. Genomic resources are still very limited for chickpea, a very important food legume crop. Here, we report the design and comprehensive validation of Next Generation Sequencing transcriptome data for chickpea through microarray technology to develop a high-throughput resource for studying the expression of all the transcripts in different biological samples to help functional genomics and breeding programs. This microarray design was developed and validated jointly by Genotypic Technology Private Limited and National Institute of Plant Genome Research. First, we designed 400k probes using reads covering 35k assembled contigs and 100k singletons chickpea transcripts. The 400k chip was hybridized with DNA and RNA samples of chickpea and microarray analysis was carried out. A total of 73,922 probes were found to be specific to chickpea transcripts. Best probes were filtered from the analyzed data and a total of 61,659 probes were selected to develop the final microarray design in 60k gene-expression microarray format. The probes represented 51,444 unique transcripts. The probes were annotated based on their corresponding chickpea transcript and similarity with other plants species. Microarray results were concordant with previous results from the NGS studies. The design of custom oligonucleotide probes for microarrays have varied functional genomic applications and this approach represents a valuable resource for chickpea. Overall design: Organism: Chickpea (Cicer arietinum) Custom Agilent 2x 400k Microarray (AMADID: 35301) designed by Genotypic Technology Pvt.Ltd.

基因芯片（Microarray）已日益成为高通量基因表达研究与新型生物标志物基因发现的强力工具。该技术已针对包括植物在内的众多生物开发完成，通常用于具备测序数据的物种，以开展基因表达分析、microRNA（miRNA）表达谱分析、比较基因组杂交（comparative genomic hybridization, CGH）、染色质免疫沉淀芯片（ChIP-on-chip）及单核苷酸多态性（SNP）分析。作为一种极为重要的食用豆科作物，鹰嘴豆的基因组学资源仍十分匮乏。本研究报道了依托基因芯片技术对鹰嘴豆二代测序（Next Generation Sequencing, NGS）转录组数据进行的设计与全面验证，旨在开发一种高通量资源，用于研究不同生物样本中所有转录本的表达情况，以助力功能基因组学与育种相关研究。本基因芯片的设计与联合验证工作由Genotypic Technology Private Limited与国家植物基因组研究所共同完成。研究团队首先利用覆盖35k个组装重叠群（contig）与100k个鹰嘴豆单例序列转录本（singleton）的测序读段，设计了400k个探针。将该400k芯片与鹰嘴豆的DNA及RNA样本进行杂交，并开展基因芯片分析。最终共有73922个探针被证实可特异性结合鹰嘴豆转录本。从分析数据中筛选出最优探针，最终选取61659个探针，以60k基因表达芯片格式构建最终的芯片设计方案。该探针组可覆盖51444个独特转录本。研究人员基于探针对应的鹰嘴豆转录本及其与其他植物物种的序列相似性对探针进行了注释。基因芯片分析结果与此前二代测序研究的结果具有良好一致性。定制寡核苷酸基因芯片探针的设计可应用于多种功能基因组学研究场景，该方案为鹰嘴豆相关研究提供了极具价值的资源。整体设计如下：实验物种：鹰嘴豆（Cicer arietinum）；由Genotypic Technology Pvt.Ltd.定制开发的安捷伦2×400k基因芯片（阵列编号：AMADID: 35301）