RNA sequencing is a prevailing research tool for transcriptomics research, which uses next-generation sequencing technology to provide information about the transcription of an organism rapidly and accurately, revealing the gene expression of an organism. In recent years, the method has become very popular in biological sciences and medical research, and its use is gradually moving towards clinical applications.
Introduction to Dual RNA Sequencing
The transcriptome is the collection of all RNAs transcribed from a specific cell or tissue at a specific time or state. RNA sequencing is a prevailing research tool for transcriptomics research, which uses next-generation sequencing technology to provide information about the transcription of an organism rapidly and accurately, revealing the gene expression of an organism. In recent years, this method has become very popular in biological sciences and medical research, and its use is gradually moving towards clinical applications.
When studying the events as bacteria infect the host, most focus on either bacterial or host mRNA. Dual RNA sequencing technology allows both to be focused simultaneously to obtain a more complete picture of the events that occur during the early and late stages of infection. It labels the bacteria with fluorescence, enabling the identification of the infected host cell components. This technique allows the study of mRNAs and transcripts of non-coding RNAs from both organisms as the course of the infection progresses.
Workflow of Dual RNA Sequencing
The parallel transcriptomic analysis of bacterial pathogens and their eukaryotic host cells is known as dual RNA-Seq. A full dual RNA-Seq test comprises three phases: (1) total host-bacteria RNA extraction and purification; (2) total RNA next-generation sequencing (NGS); and (3) bioinformatic processing and statistical analysis of the host-bacteria transcriptome.
Dual RNA-Seq entails a standard workflow that includes RNA extraction, rRNA depletion, library preparation, sequencing, and data analysis and interpretation. Meanwhile, data analysis procedures include preprocessing, alignment, and advanced downstream analyses such as alternative and non-linear splicing, differential expression, and epigenetic analyses, all of which lead to quality reports, novel transcript calculations and predictions, differentially expressed transcript probabilities, and transcript characterization.
Like traditional transcriptomic methodologies, dual RNA-seq entails quantification, difference screening, pathway annotation, network construction, and so on.
Applications of Dual RNA Sequencing
By extracting the total RNA from infected cells, dual RNA-Seq can reveal the interaction between pathogens and their hosts. While the mixed sequencing reads are allocated to their originating genomes, the sequencing and analyses of the interested RNAs from bacterial pathogens and infected cells can be accompanied at the same time, benefited from the advancement of library construction and sequencing technologies. Its use can be applied to a variety of research directions, including but not limited to:
Human Microbiome Research Promotes Progress of Precision Medicine
A large part of bacterial, archaeal, viral, and fungal microbial taxa consists of the human microbiome. While many of these microorganisms are commensal, some are harmful to humans and many are symbiotic. Our livelihoods are strongly intertwined with the microbes we allocate our bodies with regardless of whether their appearance is advantageous, insignificant, or harmful. Over the past few years, human microbiome research, defined as the research of the whole DNA component of micro-organisms that reside in our bodies, has grown quickly.
Sequencing-Based Methods and Techniques for Microbial Identification
Microbial detection and identification techniques have evolved over time, from conventional methods that are based heavily on culture, morphology, physiology, pathology, and biochemical testing, which are time-consuming and labour-intensive, to advanced mass spectrometry and genetic technologies, which have resulted in the recent surge in microbiology studies.
Fungal Identification by Sequencing: LSU, SSU, and ITS Regions
The mycobiome plays a vital role in processes such as decomposition, symbiosis, and disease progression. Knowing the role of different fungi in these processes is critical to understanding microbial communities and their impacts on environment. In drug discovery, accurate fungal identification unlocks information about species and biochemical properties.
