Related Experiment Video
Updated: Oct 17, 2025
11:26
Sequencing of mRNA from Whole Blood using Nanopore Sequencing
Published on: June 3, 2019
14.0K
Direct Nanopore Sequencing of Individual Full Length tRNA Strands
Niki K Thomas1, Vinay C Poodari1, Miten Jain1
1Biomolecular Engineering Department, Genomics Institute, and Center for Molecular Biology of RNA University of California, Santa Cruz, California 95064, United States.
ACS Nano
|October 7, 2021
Summary
Directly sequencing transfer RNA (tRNA) using nanopore technology achieves high accuracy for full-length molecules. This method identifies tRNA isoacceptors and modifications, paving the way for clinical applications.
Area of Science:
- Molecular Biology
- Genomics
- Biotechnology
Background:
- Transfer RNA (tRNA) molecules are crucial for protein synthesis, playing a vital role in translating genetic information.
- Accurate sequencing of tRNA is essential for understanding gene expression regulation and identifying disease-related variations.
- Current tRNA sequencing methods face challenges in achieving full-length molecule analysis and detecting modifications.
Purpose of the Study:
- To develop and validate a direct, high-precision sequencing method for individual transfer RNA (tRNA) molecules using Oxford Nanopore MinION technology.
- To assess the capability of nanopore sequencing in identifying tRNA isoacceptors, isodecoders, and nucleotide modifications.
- To evaluate the potential of this novel tRNA sequencing approach for future clinical diagnostics.
Main Methods:
- Utilized custom adapters for end-to-end sequencing of single tRNA molecules with subnanometer precision.
- Developed and optimized a specialized nanopore sequencing pipeline tailored for tRNA analysis.
- Tested the method on purified E. coli tRNAfMet, tRNALys, and tRNAPhe, as well as total E. coli MRE600 tRNA.
Main Results:
- Achieved 76-92% full-length sequence reads for individual aligned tRNA molecules.
- Successfully identified all 43 expected isoacceptors and isodecoders within total E. coli tRNA.
- Detected systematic nucleotide miscalls diagnostic of known tRNA modifications, including pseudouridine in the T loop.
Conclusions:
- Direct nanopore sequencing offers a promising high-throughput method for analyzing full-length tRNA molecules.
- The method accurately identifies tRNA populations and reveals nucleotide modifications, offering insights into tRNA biology.
- Further optimization is required to fully implement direct tRNA nanopore sequencing for human healthcare applications.
Related Concept Videos
03:21RNA-seq
10.5K
RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases.
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while...
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while...
10.5K
03:00Next-generation Sequencing
94.2K
The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features....
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features....
94.2K
01:57Sanger Sequencing
761.7K
DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...
761.7K