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Concepts of Transcriptomics - 20-8-2024

Concepts of Transcriptomics_20-8-2024

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7 views6 pages

Concepts of Transcriptomics - 20-8-2024

Concepts of Transcriptomics_20-8-2024

Uploaded by

ajays162616
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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20-08-2024

Concepts of Transcriptomics
20-8-2024
Types of RNAs and the respective roles in cells – microRNAs and non
coding RNAs
Overview on mRNA Seq. vs other RNA sequencing

Introduction
• Ribonucleic acid, a nucleic acid present in all living
cells.
• Its principal role is to act as a messenger carrying
instructions from DNA for controlling the synthesis of
proteins, although in some viruses (HIV) RNA rather
than DNA carries the genetic information.
• RNA is single-stranded.
• Nitrogenous bases are Adenine, Guanine, Uracil and
Cytosine.

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• Transcriptome: A transcriptome is the full range of messenger RNA, or


mRNA molecules expressed by an organism.
→Transcriptomics is the study of all RNA molecules in an organism,
using high-throughput methods.

Types of RNA
• Three main types of RNA are involved in protein synthesis.
• messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA).
• Other functions include RNA editing, gene regulation, and RNA
interference.
• Regulatory RNAs: small nuclear RNA, microRNA, and small interfering
RNA.

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Types of RNA within a typical


eukaryotic cell

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• Small nuclear RNAs (snRNAs) are non-coding RNA molecules that are
abundant in the nucleus of eukaryotic cells. They play important roles in
splicing introns from primary genomic transcripts, and are also involved
in other RNA processing events.
• MicroRNA (miRNA) is a family of small, non-coding RNA molecules that
regulate gene expression in plants, animals, and some viruses. They are
involved in many biological processes, including cell proliferation,
differentiation, apoptosis, immune response, growth, and
development.
• Small interfering RNA (siRNA), also known as short interfering RNA
or silencing RNA, are non-coding RNA molecules, belonging to a class of
double-stranded RNA molecules (20–25 bp length) and interferes with
the expression of specific genes with complementary nucleotide
sequences.
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RNA sequencing
• RNA-seq (RNA-sequencing) is a technique that can analyze the amount and sequences
of RNA in a sample.
• By analyzing the transcriptome, it can determine which of the genes that are encoded in
our DNA are active and to what degree.
• Applications
• Study the total cellular content of RNAs including mRNA, rRNA and tRNA.
• RNA-seq can tell us which genes are turned on in a cell, what their level of transcription is,
and at what times they are activated or shut off.
• identify post-transcriptional modifications that occur during mRNA processing such as
polyadenylation and 5’ capping.

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Overview of RNA-Sequencing
1. First, RNA is extracted from the biological material of
choice (e.g., cells, tissues).

2. Subsets of RNA molecules are isolated using a specific


protocol, such as the poly-A selection protocol to enrich for
polyadenylated transcripts or a ribo-depletion protocol to
remove ribosomal RNAs.

3. Next, the RNA is converted to complementary DNA


(cDNA) by reverse transcription and sequencing adaptors
are ligated to the ends of the cDNA fragments.

4. Following amplification by PCR, the RNA-Seq library is


ready for sequencing.

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Other RNA-Seq methods


• Single-cell RNA-Seq : a next-generation sequencing (NGS) method that
examines the RNA molecules in each cell of a sample. It provides a
snapshot of the transcriptome, which is the genes that were being
transcribed when the cells were harvested. This allows for a more
detailed analysis of cellular diversity and provides a high-resolution view
of cell-to-cell variation.
• CITE-Seq: Cellular Indexing of Transcriptomes and Epitopes is a
sequencing-based method that simultaneously quantifies cell surface
protein and transcriptomic data within a single cell readout.
• Spatial transcriptomics: Spatial transcriptomics is a RNA sequencing
technology where the mRNA expression is accurately resolved at cellular
level in structurally preserved tissues.

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