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Mass Spectrometry: Carboxylic Acid, Ester, and Amide Fragmentation01:01

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The fragmentation patterns observed for compounds such as carboxylic acids, esters, and amides in the mass spectra include ⍺-cleavage and McLafferty rearrangement. Fragmentation by ⍺-cleavage preferentially occurs at the carbon-carbon bond at the ⍺-position next to the carboxylic group to generate a neutral radical and a cation. Long chain compounds with hydrogen at their γ-carbon undergo McLafferty rearrangement to give a radical cation and a neutral alkene.
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Mass Spectrometry: Molecular Fragmentation Overview01:20

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The ionization of a molecule into a molecular ion inside the mass spectrometer causes instability in the molecule's structure due to the loss of an electron. This eventually leads to the fragmentation or breaking of some bonds in the molecule. The fragmentation occurs predominantly at specific bonds to yield relatively stable fragments.
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Mass Spectrometry: Amine Fragmentation00:55

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Amines can be identified using mass spectroscopy based on their characteristic fragmentation patterns. The molecular ions of amines undergo fragmentation via ⍺-cleavage. The ⍺-cleavage of the carbon-carbon bonds in amines generates an alkyl radical and resonance-stabilized nitrogen-containing cation.
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Mass Spectrometry: Long-Chain Alkane Fragmentation01:18

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RNA-seq03:21

RNA-seq

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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. 
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Mass Spectrometry: Alcohol Fragmentation01:03

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Alcohols (R-OH) ionize to lose one non-bonded electron from the oxygen atom, forming molecular ions. Due to their tendency to fragment rapidly, the intensity of the molecular ion peak in the mass spectrum is weak or sometimes absent. The fragmentation patterns for alcohols occur in two ways, i.e. ⍺-cleavage and dehydration. During ⍺-cleavage, the bond at the ⍺-position adjacent to the hydroxyl group cleaves to give a resonance-stabilized cation and a radical. However,...
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RNA Secondary Structure Prediction Using High-throughput SHAPE
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mRNA Fragmentation Pattern Detected by SHAPE.

Shanshan Feng1, Ting Chen1, Yunlong Zhang1

  • 1College of Biological Science and Medical Engineering, Donghua University, Shanghai 201620, China.

Current Issues in Molecular Biology
|September 27, 2024
PubMed
Summary
This summary is machine-generated.

Messenger RNA (mRNA) quality control is improved with multi-primer reverse transcription sequencing (MPRT-seq). This high-resolution method identifies degradation fragments, enhancing mRNA vaccine stability.

Keywords:
degradationfreeze–thaw cycleslong-term storagemRNA stabilitymRNA structure

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Vaccine Development

Background:

  • Messenger RNA (mRNA) vaccines have shown significant success, particularly against COVID-19.
  • Current quality control for mRNA relies on low-resolution electrophoresis, limiting detailed analysis of degradation.

Purpose of the Study:

  • To introduce a high-resolution method, multi-primer reverse transcription sequencing (MPRT-seq), for identifying mRNA degradation fragments.
  • To analyze mRNA completeness and degradation patterns at nucleotide resolution.

Main Methods:

  • Development and application of multi-primer reverse transcription sequencing (MPRT-seq).
  • Analysis of in-house-made mRNA constructs with antigens and untranslated regions (UTRs).
  • Correlation of sensitive base sequences with secondary structures.

Main Results:

  • MPRT-seq provides nucleotide-level resolution of mRNA degradation fragments.
  • Specific sequences at the 5' of bulge-stem-loop structures were identified as sites of preferential chain breaks.
  • Results align with capillary electrophoresis (CE) but offer significantly higher resolution.

Conclusions:

  • MPRT-seq enhances the understanding of mRNA degradation patterns.
  • Identifying sequence-structure vulnerabilities can guide the design of more stable mRNA molecules.
  • This method can improve the quality control and stability of mRNA-based therapeutics and vaccines.