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LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method
Transfer RNAs (tRNAs) are the most heavily modified RNA species in life entities. Post-transcriptional modifications severely impact the structure and function of tRNAs. To date, hundreds of modifications have been identified in tRNAs, mainly from microorganisms and animals. However, tRNAs in plant...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7226401/ https://www.ncbi.nlm.nih.gov/pubmed/32316488 http://dx.doi.org/10.3390/biom10040621 |
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author | Yan, Tongmeng Hu, Kua Ren, Fei Jiang, Zhihong |
author_facet | Yan, Tongmeng Hu, Kua Ren, Fei Jiang, Zhihong |
author_sort | Yan, Tongmeng |
collection | PubMed |
description | Transfer RNAs (tRNAs) are the most heavily modified RNA species in life entities. Post-transcriptional modifications severely impact the structure and function of tRNAs. To date, hundreds of modifications have been identified in tRNAs, mainly from microorganisms and animals. However, tRNAs in plant roots or tubers that have been widely used for food and medical purpose for centuries are rarely studied because isolation of RNA from plants still remains a challenge. In this paper, a polysaccharase-aided RNA isolation (PARI) method for extraction of high-quality RNA from plants containing large quantities of polysaccharides is developed. This method presents a new strategy of “digesting” polysaccharides that is completely different from the conventional method of “dissolving” the contaminants. By using this method, RNA of high integrity and purity were successfully extracted from ginseng roots because polysaccharide contaminations were removed efficiently with α-amylase digestion. Ginseng tRNAs were first sequenced by NGS and a total of 41 iso acceptors were identified. ChloroplastictRNA(Gly(GCC)) in ginseng root was purified and four modified nucleosides, including m(7)G, D, T, and Ψ, were identified by LC-MS/MS. The results also revealed that the m(7)G occurs at a novel position 18, which may be related to the deformation of D-loop. PARI is the first enzyme-assisted technique for RNA isolation from plants, which could fundamentally solve the problem of polysaccharide contaminations. By using the PARI method, more individual tRNAs could be isolated easily from polysaccharide-rich plant tissues, which would have a positive impact on the feasibility of research on structure and function of tRNA in plants. |
format | Online Article Text |
id | pubmed-7226401 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-72264012020-05-18 LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method Yan, Tongmeng Hu, Kua Ren, Fei Jiang, Zhihong Biomolecules Article Transfer RNAs (tRNAs) are the most heavily modified RNA species in life entities. Post-transcriptional modifications severely impact the structure and function of tRNAs. To date, hundreds of modifications have been identified in tRNAs, mainly from microorganisms and animals. However, tRNAs in plant roots or tubers that have been widely used for food and medical purpose for centuries are rarely studied because isolation of RNA from plants still remains a challenge. In this paper, a polysaccharase-aided RNA isolation (PARI) method for extraction of high-quality RNA from plants containing large quantities of polysaccharides is developed. This method presents a new strategy of “digesting” polysaccharides that is completely different from the conventional method of “dissolving” the contaminants. By using this method, RNA of high integrity and purity were successfully extracted from ginseng roots because polysaccharide contaminations were removed efficiently with α-amylase digestion. Ginseng tRNAs were first sequenced by NGS and a total of 41 iso acceptors were identified. ChloroplastictRNA(Gly(GCC)) in ginseng root was purified and four modified nucleosides, including m(7)G, D, T, and Ψ, were identified by LC-MS/MS. The results also revealed that the m(7)G occurs at a novel position 18, which may be related to the deformation of D-loop. PARI is the first enzyme-assisted technique for RNA isolation from plants, which could fundamentally solve the problem of polysaccharide contaminations. By using the PARI method, more individual tRNAs could be isolated easily from polysaccharide-rich plant tissues, which would have a positive impact on the feasibility of research on structure and function of tRNA in plants. MDPI 2020-04-17 /pmc/articles/PMC7226401/ /pubmed/32316488 http://dx.doi.org/10.3390/biom10040621 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Yan, Tongmeng Hu, Kua Ren, Fei Jiang, Zhihong LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method |
title | LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method |
title_full | LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method |
title_fullStr | LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method |
title_full_unstemmed | LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method |
title_short | LC-MS/MS Profiling of Post-Transcriptional Modifications in Ginseng tRNA Purified by a Polysaccharase-Aided Extraction Method |
title_sort | lc-ms/ms profiling of post-transcriptional modifications in ginseng trna purified by a polysaccharase-aided extraction method |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7226401/ https://www.ncbi.nlm.nih.gov/pubmed/32316488 http://dx.doi.org/10.3390/biom10040621 |
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