Organism-specific rRNA capture system for application in next-generation sequencing.

RNA-sequencing is a powerful tool in studying RNomics. However, the highly abundance of ribosomal RNAs (rRNA) and transfer RNA (tRNA) have predominated in the sequencing reads, thereby hindering the study of lowly expressed genes. Therefore, rRNA depletion prior to sequencing is often performed in o...

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Autores principales:	Sai-Kam Li, Jun-Wei Zhou, Aldrin Kay-Yuen Yim, Alden King-Yung Leung, Stephen Kwok-Wing Tsui, Ting-Fung Chan, Terrence Chi-Kong Lau
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/2b2c2f9761a948b1954cd32b70f6f1c2
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spelling	oai:doaj.org-article:2b2c2f9761a948b1954cd32b70f6f1c22021-11-18T08:54:25ZOrganism-specific rRNA capture system for application in next-generation sequencing.1932-620310.1371/journal.pone.0074286https://doaj.org/article/2b2c2f9761a948b1954cd32b70f6f1c22013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24073205/?tool=EBIhttps://doaj.org/toc/1932-6203RNA-sequencing is a powerful tool in studying RNomics. However, the highly abundance of ribosomal RNAs (rRNA) and transfer RNA (tRNA) have predominated in the sequencing reads, thereby hindering the study of lowly expressed genes. Therefore, rRNA depletion prior to sequencing is often performed in order to preserve the subtle alteration in gene expression especially those at relatively low expression levels. One of the commercially available methods is to use DNA or RNA probes to hybridize to the target RNAs. However, there is always a concern with the non-specific binding and unintended removal of messenger RNA (mRNA) when the same set of probes is applied to different organisms. The degree of such unintended mRNA removal varies among organisms due to organism-specific genomic variation. We developed a computer-based method to design probes to deplete rRNA in an organism-specific manner. Based on the computation results, biotinylated-RNA-probes were produced by in vitro transcription and were used to perform rRNA depletion with subtractive hybridization. We demonstrated that the designed probes of 16S rRNAs and 23S rRNAs can efficiently remove rRNAs from Mycobacterium smegmatis. In comparison with a commercial subtractive hybridization-based rRNA removal kit, using organism-specific probes is better in preserving the RNA integrity and abundance. We believe the computer-based design approach can be used as a generic method in preparing RNA of any organisms for next-generation sequencing, particularly for the transcriptome analysis of microbes.Sai-Kam LiJun-Wei ZhouAldrin Kay-Yuen YimAlden King-Yung LeungStephen Kwok-Wing TsuiTing-Fung ChanTerrence Chi-Kong LauPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 9, p e74286 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Sai-Kam Li Jun-Wei Zhou Aldrin Kay-Yuen Yim Alden King-Yung Leung Stephen Kwok-Wing Tsui Ting-Fung Chan Terrence Chi-Kong Lau Organism-specific rRNA capture system for application in next-generation sequencing.
description	RNA-sequencing is a powerful tool in studying RNomics. However, the highly abundance of ribosomal RNAs (rRNA) and transfer RNA (tRNA) have predominated in the sequencing reads, thereby hindering the study of lowly expressed genes. Therefore, rRNA depletion prior to sequencing is often performed in order to preserve the subtle alteration in gene expression especially those at relatively low expression levels. One of the commercially available methods is to use DNA or RNA probes to hybridize to the target RNAs. However, there is always a concern with the non-specific binding and unintended removal of messenger RNA (mRNA) when the same set of probes is applied to different organisms. The degree of such unintended mRNA removal varies among organisms due to organism-specific genomic variation. We developed a computer-based method to design probes to deplete rRNA in an organism-specific manner. Based on the computation results, biotinylated-RNA-probes were produced by in vitro transcription and were used to perform rRNA depletion with subtractive hybridization. We demonstrated that the designed probes of 16S rRNAs and 23S rRNAs can efficiently remove rRNAs from Mycobacterium smegmatis. In comparison with a commercial subtractive hybridization-based rRNA removal kit, using organism-specific probes is better in preserving the RNA integrity and abundance. We believe the computer-based design approach can be used as a generic method in preparing RNA of any organisms for next-generation sequencing, particularly for the transcriptome analysis of microbes.
format	article
author	Sai-Kam Li Jun-Wei Zhou Aldrin Kay-Yuen Yim Alden King-Yung Leung Stephen Kwok-Wing Tsui Ting-Fung Chan Terrence Chi-Kong Lau
author_facet	Sai-Kam Li Jun-Wei Zhou Aldrin Kay-Yuen Yim Alden King-Yung Leung Stephen Kwok-Wing Tsui Ting-Fung Chan Terrence Chi-Kong Lau
author_sort	Sai-Kam Li
title	Organism-specific rRNA capture system for application in next-generation sequencing.
title_short	Organism-specific rRNA capture system for application in next-generation sequencing.
title_full	Organism-specific rRNA capture system for application in next-generation sequencing.
title_fullStr	Organism-specific rRNA capture system for application in next-generation sequencing.
title_full_unstemmed	Organism-specific rRNA capture system for application in next-generation sequencing.
title_sort	organism-specific rrna capture system for application in next-generation sequencing.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/2b2c2f9761a948b1954cd32b70f6f1c2
work_keys_str_mv	AT saikamli organismspecificrrnacapturesystemforapplicationinnextgenerationsequencing AT junweizhou organismspecificrrnacapturesystemforapplicationinnextgenerationsequencing AT aldrinkayyuenyim organismspecificrrnacapturesystemforapplicationinnextgenerationsequencing AT aldenkingyungleung organismspecificrrnacapturesystemforapplicationinnextgenerationsequencing AT stephenkwokwingtsui organismspecificrrnacapturesystemforapplicationinnextgenerationsequencing AT tingfungchan organismspecificrrnacapturesystemforapplicationinnextgenerationsequencing AT terrencechikonglau organismspecificrrnacapturesystemforapplicationinnextgenerationsequencing
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Organism-specific rRNA capture system for application in next-generation sequencing.

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