RNAseq Experimental design: challenges and considerations for Small RNA Sequencing of an invertebrate model - P2P Teaching, Season 3 (In-Person)

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Welcome to the third season of our Peer-to-Peer Teaching sessions. These are taught by our own community members to fill knowledge gaps and keep up with the accelerated pace at which bioinformatics grows. This season we are pleased to announce sessions covering * Image analysis methodologies (imageJ and QuPath) * Omics analysis in R and/or Python * Biomedical Data analysis using the clusters * Statistical analysis software (Prism) *Science communication (Illustrator).

This session is on "RNAseq Experimental design: challenges and considerations for Small RNA Sequencing of an invertebrate model". This lecture was designed and will be delivered by Rafael Pimenta, PhD, from the Aksoy Lab, Department of Epidemiology of Microbial Diseases in the Yale School of Public Health.

Summary:

In this lecture, we will discuss the initial steps involved in microRNA next-generation sequencing, bearing in mind special considerations of working with invertebrate samples as a template. In the Diptera order, ribosomal RNA is over-abundant and can make up 95% of all small RNA reads, and unhindered amplification of 2S rRNA greatly impairs output analysis by overwhelming the sequencing process. Nevertheless, preparing biological samples for miRNA-seq is a relatively simple process, as several protocols adaptations have been designed that can efficiently remove 2S rRNA.

This workshop is designed for:

Beginner-level students and researchers interested in:

• Understanding experimental design of RNA-seq methods.
• Invertebrate-oriented researchers curious about 2S RNA elimination from RNA-seq samples.
• Beginner-level introduction of FASTQ analyzes.

In this Lecture You will learn about:

1. Setting up miRNA-seq Workflow:
   1. Overview of QIAseq miRNA Library prep protocol
   2. Addition of modified step for 2S rRNA removal
2. Analysis of the Sequencing Data:
   1. FASTQC: check miRNA reads quality (verify 2S removal):
   2. CLC Genomic Workbench pipeline (quantification and alignment of reads)
   3. Unmapped reads? How to identify reads to other reference genome.

Requirements:

This lecture is a demonstration. There are no requirements to participate in this lecture. Your presence and interest are sufficient!