miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.

<h4>Background</h4>One of the most widely used evaluation methods in miRNA experiments is qRT-PCR. However, selecting suitable internal controls (IC) is crucial for qRT-PCR experiments. Currently, there is no consensus on the ICs for miRNA qRT-PCR experiments in breast cancer. To this en...

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Autores principales:	Sogol Ghanbari, Adel Salimi, Saeid Rahmani, Nahid Nafissi, Ali Sharifi-Zarchi, Seyed Javad Mowla
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/75b4965c643b47c7a4e3ab2f9f8123ad
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spelling	oai:doaj.org-article:75b4965c643b47c7a4e3ab2f9f8123ad2021-12-02T20:10:59ZmiR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.1932-620310.1371/journal.pone.0253009https://doaj.org/article/75b4965c643b47c7a4e3ab2f9f8123ad2021-01-01T00:00:00Zhttps://doi.org/10.1371/journal.pone.0253009https://doaj.org/toc/1932-6203<h4>Background</h4>One of the most widely used evaluation methods in miRNA experiments is qRT-PCR. However, selecting suitable internal controls (IC) is crucial for qRT-PCR experiments. Currently, there is no consensus on the ICs for miRNA qRT-PCR experiments in breast cancer. To this end, we tried to identify the most stable (the least expression alteration) and promising miRNAs in normal and tumor breast tissues by employing TCGA miRNA-Seq data and then experimentally validated them on fresh clinical samples.<h4>Methods</h4>A multi-component scoring system was used which takes into account multiple expression stability criteria as well as correlation with clinical characteristics. Furthermore, we extended the scoring system for more than two biological sub-groups. TCGA BRCA samples were analyzed based on two grouping criteria: Tumor & Normal samples and Tumor subtypes. The top 10 most stable miRNAs were further investigated by differential expression and survival analysis. Then, we examined the expression level of the top scored miRNA (hsa-miR-361-5p) along with two commonly used ICs hsa-miR-16-5p and U48 on 34 pairs of Primary breast tumor and their adjacent normal tissues using qRT-PCR.<h4>Results</h4>According to our multi-component scoring system, hsa-miR-361-5p had the highest stability score in both grouping criteria and hsa-miR-16-5p showed significantly lower scores. Based on our qRT-PCR assay, while U48 was the most abundant IC, hsa-miR-361-5p had lower standard deviation and also was the only IC capable of detecting a significant up-regulation of hsa-miR-21-5p in breast tumor tissue.<h4>Conclusions</h4>miRNA-Seq data is a great source to discover stable ICs. Our results demonstrated that hsa-miR-361-5p is a highly stable miRNA in tumor and non-tumor breast tissue and we recommend it as a suitable reference gene for miRNA expression studies in breast cancer. Additionally, although hsa-miR-16-5p is a commonly used IC, it's not a suitable one for breast cancer studies.Sogol GhanbariAdel SalimiSaeid RahmaniNahid NafissiAli Sharifi-ZarchiSeyed Javad MowlaPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 16, Iss 6, p e0253009 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Sogol Ghanbari Adel Salimi Saeid Rahmani Nahid Nafissi Ali Sharifi-Zarchi Seyed Javad Mowla miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.
description	<h4>Background</h4>One of the most widely used evaluation methods in miRNA experiments is qRT-PCR. However, selecting suitable internal controls (IC) is crucial for qRT-PCR experiments. Currently, there is no consensus on the ICs for miRNA qRT-PCR experiments in breast cancer. To this end, we tried to identify the most stable (the least expression alteration) and promising miRNAs in normal and tumor breast tissues by employing TCGA miRNA-Seq data and then experimentally validated them on fresh clinical samples.<h4>Methods</h4>A multi-component scoring system was used which takes into account multiple expression stability criteria as well as correlation with clinical characteristics. Furthermore, we extended the scoring system for more than two biological sub-groups. TCGA BRCA samples were analyzed based on two grouping criteria: Tumor & Normal samples and Tumor subtypes. The top 10 most stable miRNAs were further investigated by differential expression and survival analysis. Then, we examined the expression level of the top scored miRNA (hsa-miR-361-5p) along with two commonly used ICs hsa-miR-16-5p and U48 on 34 pairs of Primary breast tumor and their adjacent normal tissues using qRT-PCR.<h4>Results</h4>According to our multi-component scoring system, hsa-miR-361-5p had the highest stability score in both grouping criteria and hsa-miR-16-5p showed significantly lower scores. Based on our qRT-PCR assay, while U48 was the most abundant IC, hsa-miR-361-5p had lower standard deviation and also was the only IC capable of detecting a significant up-regulation of hsa-miR-21-5p in breast tumor tissue.<h4>Conclusions</h4>miRNA-Seq data is a great source to discover stable ICs. Our results demonstrated that hsa-miR-361-5p is a highly stable miRNA in tumor and non-tumor breast tissue and we recommend it as a suitable reference gene for miRNA expression studies in breast cancer. Additionally, although hsa-miR-16-5p is a commonly used IC, it's not a suitable one for breast cancer studies.
format	article
author	Sogol Ghanbari Adel Salimi Saeid Rahmani Nahid Nafissi Ali Sharifi-Zarchi Seyed Javad Mowla
author_facet	Sogol Ghanbari Adel Salimi Saeid Rahmani Nahid Nafissi Ali Sharifi-Zarchi Seyed Javad Mowla
author_sort	Sogol Ghanbari
title	miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.
title_short	miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.
title_full	miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.
title_fullStr	miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.
title_full_unstemmed	miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.
title_sort	mir-361-5p as a promising qrt-pcr internal control for tumor and normal breast tissues.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/75b4965c643b47c7a4e3ab2f9f8123ad
work_keys_str_mv	AT sogolghanbari mir3615pasapromisingqrtpcrinternalcontrolfortumorandnormalbreasttissues AT adelsalimi mir3615pasapromisingqrtpcrinternalcontrolfortumorandnormalbreasttissues AT saeidrahmani mir3615pasapromisingqrtpcrinternalcontrolfortumorandnormalbreasttissues AT nahidnafissi mir3615pasapromisingqrtpcrinternalcontrolfortumorandnormalbreasttissues AT alisharifizarchi mir3615pasapromisingqrtpcrinternalcontrolfortumorandnormalbreasttissues AT seyedjavadmowla mir3615pasapromisingqrtpcrinternalcontrolfortumorandnormalbreasttissues
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miR-361-5p as a promising qRT-PCR internal control for tumor and normal breast tissues.

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