In addition to the canonical nucleobases, a variety of chemical modifications have been identified presence in nucleic acids. These modifications have been demonstrated to involve in regulating the spatiotemporal expr...In addition to the canonical nucleobases, a variety of chemical modifications have been identified presence in nucleic acids. These modifications have been demonstrated to involve in regulating the spatiotemporal expression of genes. Up to date, over 150 types of chemical modifications have been found existence in nucleic acids. Understanding the functional roles of modifications relies on deciphering the location information of modifications in nucleic acids. Analytical methods for studying nucleic acid modifications have greatly advanced in the last decade. To locate the modifications in nucleic acids, various mass spectrometry (MS)-based analytical strategies have been established. Recent progress in next-generation sequencing (NGS) in conjugation with immunoprecipitation, chemical reaction, enzyme-mediated mutation, or nanomaterials offer genome-wide or transcriptome-wide mapping of modifications, which greatly revolutionize the field of epigenetic modifications. Herein, we reviewed and summarized the established methods and the breakthrough of the techniques for locating modifications in nucleic acids. In addition, we discussed the principles, applications, advantages and drawbacks of these methods. We believe that with the rapid advancement of techniques and methods,the functions of nucleic acid modifications will be fully understood in the future.展开更多
RNA modification has recently been proposed to play important roles in biological regulation. The detection and quantification of RNA modifications generally are challenging tasks since most of the modifications exist...RNA modification has recently been proposed to play important roles in biological regulation. The detection and quantification of RNA modifications generally are challenging tasks since most of the modifications exist in low abundance in vivo. Here we developed an on-line trapping/capillary hydrophilic-interaction liquid chromatography/electrospray ionization-mass spectrometry(on-line trapping/cHILIC/MS) method for sensitive and simultaneous quantification of RNA modifications of N^6-methyladenosine(m^6A) and 5-methylcytosine(5-mC) from human blood. The hydrophilic organic-silica hybrid monolith was prepared using sol-gel combined with "thiol-ene" click reaction for the separation of nucleosides. A poly(MAA-co-EGDMA) monolithic capillary was used as the on-line trapping column.With the developed on-line trapping/cHILIC/MS analytical platform, the detection limits of m^6A and 5-mC can reach to 0.06 fmol and 0.10 fmol. We then investigated the contents of m^6A and 5-mC in human blood RNA from healthy persons at the age of 6-14 and 60-68 years. Our results showed that both m^6A and 5-mC contents were significantly decreased in elder persons, suggesting the RNA modifications of m^6A and 5-mC are correlated to aging.展开更多
基金the National Key R&D Program of China (No. 2017YFC0906800)the National Natural Science Foundation of China (Nos. 21672166, 21635006, 21721005, 21728802) for the financial support
文摘In addition to the canonical nucleobases, a variety of chemical modifications have been identified presence in nucleic acids. These modifications have been demonstrated to involve in regulating the spatiotemporal expression of genes. Up to date, over 150 types of chemical modifications have been found existence in nucleic acids. Understanding the functional roles of modifications relies on deciphering the location information of modifications in nucleic acids. Analytical methods for studying nucleic acid modifications have greatly advanced in the last decade. To locate the modifications in nucleic acids, various mass spectrometry (MS)-based analytical strategies have been established. Recent progress in next-generation sequencing (NGS) in conjugation with immunoprecipitation, chemical reaction, enzyme-mediated mutation, or nanomaterials offer genome-wide or transcriptome-wide mapping of modifications, which greatly revolutionize the field of epigenetic modifications. Herein, we reviewed and summarized the established methods and the breakthrough of the techniques for locating modifications in nucleic acids. In addition, we discussed the principles, applications, advantages and drawbacks of these methods. We believe that with the rapid advancement of techniques and methods,the functions of nucleic acid modifications will be fully understood in the future.
基金financially supported by the National Natural Science Foundation of China (Nos. 21522507, 21672166, 21728802, 21721005)
文摘RNA modification has recently been proposed to play important roles in biological regulation. The detection and quantification of RNA modifications generally are challenging tasks since most of the modifications exist in low abundance in vivo. Here we developed an on-line trapping/capillary hydrophilic-interaction liquid chromatography/electrospray ionization-mass spectrometry(on-line trapping/cHILIC/MS) method for sensitive and simultaneous quantification of RNA modifications of N^6-methyladenosine(m^6A) and 5-methylcytosine(5-mC) from human blood. The hydrophilic organic-silica hybrid monolith was prepared using sol-gel combined with "thiol-ene" click reaction for the separation of nucleosides. A poly(MAA-co-EGDMA) monolithic capillary was used as the on-line trapping column.With the developed on-line trapping/cHILIC/MS analytical platform, the detection limits of m^6A and 5-mC can reach to 0.06 fmol and 0.10 fmol. We then investigated the contents of m^6A and 5-mC in human blood RNA from healthy persons at the age of 6-14 and 60-68 years. Our results showed that both m^6A and 5-mC contents were significantly decreased in elder persons, suggesting the RNA modifications of m^6A and 5-mC are correlated to aging.
