RNA structure probing using long-read nanopore sequencing technology
RNA can fold into complex structures that can perform specific biological functions and the ability to characterise the structures of the RNA is the key in understanding its functions. Therefore, various groups have developed several methods that utilizes enzymatic or chemical structure probes an...
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| Other Authors: | |
| Format: | Thesis-Doctor of Philosophy |
| Language: | English |
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Nanyang Technological University
2022
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| Online Access: | https://hdl.handle.net/10356/158914 |
| _version_ | 1826123472186638336 |
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| author | Aw, Ashley Jong Ghut |
| author2 | Francesc Xavier Roca Castella |
| author_facet | Francesc Xavier Roca Castella Aw, Ashley Jong Ghut |
| author_sort | Aw, Ashley Jong Ghut |
| collection | NTU |
| description | RNA can fold into complex structures that can perform specific biological functions
and the ability to characterise the structures of the RNA is the key in understanding
its functions. Therefore, various groups have developed several methods that utilizes
enzymatic or chemical structure probes and coupling with high-throughput short-read
sequencing to obtain the structural information from in-vitro and in-vivo samples.
However, high-throughput short-read sequencing has its limitation, where it is difficult
to accurately generate the structural information for gene with multiple isoforms and
the structural information between reads are lost. Therefore, we developed POREcupine, a method that uses direct RNA sequencing, a high-throughput long-read
sequencing technology to detect the modifications caused by NAI-N3, a singlestranded chemical probe. Using PORE-cupine, we showed that shared sequences in
different transcript isoforms of the same gene can fold into different structures. We
also demonstrate that structural differences between transcript isoforms of the same
gene lead to differences in translation efficiency. We also apply PORE-cupine to
investigate the structural difference of the RNA of the wild type and Δ382 strain of
SARS-CoV2. Similarly, we found that shared sequences in different subgenomic
RNA shows structural differences, highlighting the importance of long-read
sequencing for obtaining phase information. Lastly, we have shown that besides
NAI-N3, PORE-cupine can detect other structure probing compounds, like 1AI and
CMCT. |
| first_indexed | 2024-10-01T06:04:56Z |
| format | Thesis-Doctor of Philosophy |
| id | ntu-10356/158914 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T06:04:56Z |
| publishDate | 2022 |
| publisher | Nanyang Technological University |
| record_format | dspace |
| spelling | ntu-10356/1589142023-02-28T18:42:37Z RNA structure probing using long-read nanopore sequencing technology Aw, Ashley Jong Ghut Francesc Xavier Roca Castella School of Biological Sciences Genome Institute of Singapore Wan Yue xroca@ntu.edu.sg Science::Biological sciences RNA can fold into complex structures that can perform specific biological functions and the ability to characterise the structures of the RNA is the key in understanding its functions. Therefore, various groups have developed several methods that utilizes enzymatic or chemical structure probes and coupling with high-throughput short-read sequencing to obtain the structural information from in-vitro and in-vivo samples. However, high-throughput short-read sequencing has its limitation, where it is difficult to accurately generate the structural information for gene with multiple isoforms and the structural information between reads are lost. Therefore, we developed POREcupine, a method that uses direct RNA sequencing, a high-throughput long-read sequencing technology to detect the modifications caused by NAI-N3, a singlestranded chemical probe. Using PORE-cupine, we showed that shared sequences in different transcript isoforms of the same gene can fold into different structures. We also demonstrate that structural differences between transcript isoforms of the same gene lead to differences in translation efficiency. We also apply PORE-cupine to investigate the structural difference of the RNA of the wild type and Δ382 strain of SARS-CoV2. Similarly, we found that shared sequences in different subgenomic RNA shows structural differences, highlighting the importance of long-read sequencing for obtaining phase information. Lastly, we have shown that besides NAI-N3, PORE-cupine can detect other structure probing compounds, like 1AI and CMCT. Doctor of Philosophy 2022-06-01T12:17:15Z 2022-06-01T12:17:15Z 2022 Thesis-Doctor of Philosophy Aw, A. J. G. (2022). RNA structure probing using long-read nanopore sequencing technology. Doctoral thesis, Nanyang Technological University, Singapore. https://hdl.handle.net/10356/158914 https://hdl.handle.net/10356/158914 en This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License (CC BY-NC 4.0). application/pdf Nanyang Technological University |
| spellingShingle | Science::Biological sciences Aw, Ashley Jong Ghut RNA structure probing using long-read nanopore sequencing technology |
| title | RNA structure probing using long-read nanopore sequencing technology |
| title_full | RNA structure probing using long-read nanopore sequencing technology |
| title_fullStr | RNA structure probing using long-read nanopore sequencing technology |
| title_full_unstemmed | RNA structure probing using long-read nanopore sequencing technology |
| title_short | RNA structure probing using long-read nanopore sequencing technology |
| title_sort | rna structure probing using long read nanopore sequencing technology |
| topic | Science::Biological sciences |
| url | https://hdl.handle.net/10356/158914 |
| work_keys_str_mv | AT awashleyjongghut rnastructureprobingusinglongreadnanoporesequencingtechnology |