Related Experiment Video
Updated: Nov 11, 2025
12:01
3' End Sequencing Library Preparation with A-seq2
Published on: October 10, 2017
10.9K
SPAAC-NAD-seq, a sensitive and accurate method to profile NAD+-capped transcripts.
Hao Hu1,2, Nora Flynn1, Hailei Zhang3
1Department of Botany and Plant Sciences, Institute of Integrative Genome Biology, University of California, Riverside, CA 92521.
Summary
Nicotinamide adenine diphosphate (NAD+) caps RNA in various organisms. A new method, SPAAC-NAD-seq, specifically identifies these NAD-RNAs, improving accuracy and retaining full-length sequence information.
Area of Science:
- Molecular Biology
- Genomics
- RNA Biology
Background:
- Nicotinamide adenine diphosphate (NAD+) functions as a novel RNA 5' cap in prokaryotes and eukaryotes.
- Existing methods like NAD captureSeq have limitations in specificity and RNA integrity.
Purpose of the Study:
- To develop a more specific and efficient method for identifying NAD+-capped RNAs (NAD-RNAs).
- To overcome the limitations of previous techniques regarding specificity and RNA fragmentation.
Main Methods:
- Development of a copper-free, strain-promoted azide-alkyne cycloaddition (SPAAC) based method for NAD-RNA capture.
- Comparative analysis of CuAAC and SPAAC specificity for NAD-RNAs versus m7G-capped RNAs (m7G-RNAs).
- Combination of SPAAC with immunodepletion of m7G-RNAs for enhanced NAD-RNA identification.
Main Results:
- Both CuAAC and SPAAC reactions show preference for NAD-RNAs but also react with m7G-RNAs.
- SPAAC-NAD sequencing, coupled with m7G-RNA depletion, accurately and sensitively identifies NAD-RNAs.
- New NAD-RNA profiles were discovered in Arabidopsis using the improved method.
- SPAAC-NAD-seq preserves full-length RNA sequence information.
Conclusions:
- SPAAC-NAD-seq offers a specific and efficient approach for discovering NAD-RNAs.
- This method is suitable for both prokaryotic and eukaryotic systems, especially when combined with m7G-RNA depletion.
Related Concept Videos
03:21RNA-seq
10.9K
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...
10.9K
02:24Ribosome Profiling
3.8K
Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique...
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique...
3.8K