Genome of an organism is its blueprint and sequencing is the first step in understanding it. In order to derive meaning out of the sequence data, annotation is performed. These processes collectively make DNA sequences a knowledge base for all genetic information. Fred Sanger invented a DNA sequencing method in the late 70’s that remains till date, the gold standard for individual genes.
Whole Genome Bisulphite Sequencing (WGBS) is an NGS based genome-wide methylation profiling application. Its application comprises identification and quantification of the methylation status of the entire genome. Epigenetic variations such as DNA methylation and histone modification account for differential expression of genes.
Genotyping requires thousands of genomes to be compared in a reliable, consistent way. Restriction site associated DNA sequencing (RAD-Seq) interrogates a fraction of the genome across many individuals, an ideal method for genotyping.
1. By using restriction enzyme digestion and sequencing the regions adjacent to restriction sites, researchers can examine the same subset of genomic regions for thousands of individuals and identify many genetic markers along the genome.
2. Other NGS methods examine a larger portion of the genome and offer more data, but they are costly and cannot be used to study the thousands of individuals required for genotyping.
The protein coding, or exonic regions, constitute a small percentage of the eukaryotic genome (for example, ~1.5% in the case of human genome). Mutations identified in the exonic regions typically contribute to affecting the function of proteins and are more actionable than variations that occur in non-coding regions. Exome sequencing involves selective capture and sequencing of these protein coding regions of the genome. Compared to whole genome sequencing, Exome sequencing is more cost effective and also provides better per base coverage.
RNA-Seq is a high-throughput sequencing based approach for transcriptome profiling. The information contained in the genome is transferred to RNA molecules by the process of transcription. The transcribed regions of the genome encode different types of RNA molecules, like mRNA, tRNA, rRNA, miRNA, lincRNA, snoRNA and others. The spectrum of transcribed RNA products constitute the transcriptome. In humans, the transcriptome represents an estimated 5% of the genome.
The advent of NGS technologies and their rapid advancements have revolutionized genomics research. Among them, RNA sequencing has emerged as a cost-effective means of rapidly acquiring functional sequence information for many organisms. RNA-Seq is a powerful tool for simultaneous transcriptome characterization and differential gene expression (DGE) analysis in a single cell type, tissue type, or entire organism under defined conditions. RNA-Seq has helped in decoding transcriptomes of various species, including several plants, birds, insects and animals for which no reference genome is available. RNA-Seq also provides data about alternative splicing, strand-specific expression, expression of unannotated exons (or genes), and identification of fusion genes in cancer. It also provides gene expression data at a greater sensitivity than microarrays.
NextGen LIMS is an end-to-end work flow management system for next-gen sequencing labs. It provides an automated work flow for key functions within a next-gen lab: Project Management, Sample Management, Library Management and Sequencing Run Management. It tracks the end-to-end project progression - samples, libraries and sequencing runs through a set of pre-defined workflows; thus reducing manual intervention.