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Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR

The brown marmorated stink bug (BMSB) is native to Asia and recently invaded the USA. RNA interference (RNAi) is a gene silencing mechanism in which the introduction of double-stranded RNA (dsRNA) inhibits gene function by degrading target mRNA. In dsRNA stability assays, the dsRNases present in the...

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Autores principales: Mogilicherla, Kanakachari, Howell, Jeffrey L., Palli, Subba Reddy
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5829169/
https://www.ncbi.nlm.nih.gov/pubmed/29487333
http://dx.doi.org/10.1038/s41598-018-22035-z
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author Mogilicherla, Kanakachari
Howell, Jeffrey L.
Palli, Subba Reddy
author_facet Mogilicherla, Kanakachari
Howell, Jeffrey L.
Palli, Subba Reddy
author_sort Mogilicherla, Kanakachari
collection PubMed
description The brown marmorated stink bug (BMSB) is native to Asia and recently invaded the USA. RNA interference (RNAi) is a gene silencing mechanism in which the introduction of double-stranded RNA (dsRNA) inhibits gene function by degrading target mRNA. In dsRNA stability assays, the dsRNases present in the hemolymph and salivary gland secretions of BMSB showed lower activity than those in the hemolymph of Heliothis virescens. We evaluated six housekeeping genes (18S rRNA, EF1-α, Actin, Ubiquitin, 60S RP and β-Tubulin) across dsRNA treatments (injection and feeding) in nymphs and adults of BMSB and identified 18S rRNA and 60S RP as the best genes to use as a reference in reverse-transcriptase quantitative PCR (RT-qPCR). Homologs of 13 genes that were shown to function as effective RNAi targets in other insects were identified and evaluated by injecting dsRNA targeting these homologs into BMSB adults. Five out of 13 dsRNAs tested caused more than 70% mortality by seven days after injection of dsRNA. Feeding dsRNA targeting five of these genes (IAP, ATPase, SNF7, GPCR, and PPI) to nymphs caused more than 70% mortality by three of the five dsRNAs tested. These data suggest that feeding dsRNA causes target gene knockdown and mortality in BMSB.
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spelling pubmed-58291692018-03-01 Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR Mogilicherla, Kanakachari Howell, Jeffrey L. Palli, Subba Reddy Sci Rep Article The brown marmorated stink bug (BMSB) is native to Asia and recently invaded the USA. RNA interference (RNAi) is a gene silencing mechanism in which the introduction of double-stranded RNA (dsRNA) inhibits gene function by degrading target mRNA. In dsRNA stability assays, the dsRNases present in the hemolymph and salivary gland secretions of BMSB showed lower activity than those in the hemolymph of Heliothis virescens. We evaluated six housekeeping genes (18S rRNA, EF1-α, Actin, Ubiquitin, 60S RP and β-Tubulin) across dsRNA treatments (injection and feeding) in nymphs and adults of BMSB and identified 18S rRNA and 60S RP as the best genes to use as a reference in reverse-transcriptase quantitative PCR (RT-qPCR). Homologs of 13 genes that were shown to function as effective RNAi targets in other insects were identified and evaluated by injecting dsRNA targeting these homologs into BMSB adults. Five out of 13 dsRNAs tested caused more than 70% mortality by seven days after injection of dsRNA. Feeding dsRNA targeting five of these genes (IAP, ATPase, SNF7, GPCR, and PPI) to nymphs caused more than 70% mortality by three of the five dsRNAs tested. These data suggest that feeding dsRNA causes target gene knockdown and mortality in BMSB. Nature Publishing Group UK 2018-02-27 /pmc/articles/PMC5829169/ /pubmed/29487333 http://dx.doi.org/10.1038/s41598-018-22035-z Text en © The Author(s) 2018 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Mogilicherla, Kanakachari
Howell, Jeffrey L.
Palli, Subba Reddy
Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR
title Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR
title_full Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR
title_fullStr Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR
title_full_unstemmed Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR
title_short Improving RNAi in the Brown Marmorated Stink Bug: Identification of target genes and reference genes for RT-qPCR
title_sort improving rnai in the brown marmorated stink bug: identification of target genes and reference genes for rt-qpcr
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5829169/
https://www.ncbi.nlm.nih.gov/pubmed/29487333
http://dx.doi.org/10.1038/s41598-018-22035-z
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