Cardiff University | Prifysgol Caerdydd ORCA
Online Research @ Cardiff 
WelshClear Cookie - decide language by browser settings

Quantitative expression analysis in brassica napus by northern blot analysis and reverse transcription-quantitative PCR in a complex experimental setting

Rumlow, Annekathrin, Keunen, Els, Klein, Jan, Pallmann, Philip, Riemenschneider, Anja, Cuypers, Ann and Papenbrock, Jutta 2016. Quantitative expression analysis in brassica napus by northern blot analysis and reverse transcription-quantitative PCR in a complex experimental setting. PLoS ONE 11 (9) , e0163679. 10.1371/journal.pone.0163679

Full text not available from this repository.

Abstract

Analysis of gene expression is one of the major ways to better understand plant reactions to changes in environmental conditions. The comparison of many different factors influencing plant growth challenges the gene expression analysis for specific gene-targeted experiments, especially with regard to the choice of suitable reference genes. The aim of this study is to compare expression results obtained by Northern blot, semi-quantitative PCR and RT-qPCR, and to identify a reliable set of reference genes for oilseed rape (Brassica napus L.) suitable for comparing gene expression under complex experimental conditions. We investigated the influence of several factors such as sulfur deficiency, different time points during the day, varying light conditions, and their interaction on gene expression in oilseed rape plants. The expression of selected reference genes was indeed influenced under these conditions in different ways. Therefore, a recently developed algorithm, called GrayNorm, was applied to validate a set of reference genes for normalizing results obtained by Northern blot analysis. After careful comparison of the three methods mentioned above, Northern blot analysis seems to be a reliable and cost-effective alternative for gene expression analysis under a complex growth regime. For using this method in a quantitative way a number of references was validated revealing that for our experiment a set of three references provides an appropriate normalization. Semi-quantitative PCR was prone to many handling errors and difficult to control while RT-qPCR was very sensitive to expression fluctuations of the reference genes.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Medicine
Publisher: Public Library of Science
ISSN: 1932-6203
Date of Acceptance: 11 September 2016
Last Modified: 19 Jun 2019 13:01
URI: http://orca.cf.ac.uk/id/eprint/105705

Citation Data

Cited 3 times in Scopus. View in Scopus. Powered By Scopus® Data

Actions (repository staff only)

Edit Item Edit Item