Reference genes for accessing differential expression among developmental stages and analysis of differential expression of OBP genes in Anastrepha obliqua.

Aline Minali Nakamura,Samira Chahad-Ehlers,Iderval Sobrinho,Cristiane Hayumi Taniguti,Felipe Rafael Torres,Reinaldo Alves De Brito,André Luís A Lima

doi:10.1038/srep17480

Abstract

The West Indian fruit fly, Anastrepha obliqua, is an important agricultural pest in the New World. The use of pesticide-free methods to control invasive species such as this reinforces the search for genes potentially useful in their genetic control. Therefore, the study of chemosensory proteins involved with a range of responses to the chemical environment will help not only on the understanding of the species biology but may also help the development of environmentally friendly pest control strategies. Here we analyzed the expression patterns of three OBP genes, Obp19d_2, Obp56a and Obp99c, across different phases of A. obliqua development by qPCR. In order to do so, we tested eight and identified three reference genes for data normalization, rpl17, rpl18 and ef1a, which displayed stability for the conditions here tested. All OBPs showed differential expression on adults and some differential expression among adult stages. Obp99c had an almost exclusive expression in males and Obp56a showed high expression in virgin females. Thereby, our results provide relevant data not only for other gene expression studies in this species, as well as for the search of candidate genes that may help in the development of new pest control strategies.

Highlights

Oviposition sites[6,7,8,9,10], genes that code for proteins involved in olfactory reception may prove useful in this regard as well
Normalization is an essential step for the quantitative study of relative gene expression, which means that the relative expression of a target gene must be determined against a reference gene, whose expression
Every Quantitative polymerase chain reaction (qPCR) experiment should begin with the identification and validation of a set of reference genes that can be reliably used according to a specific condition

Summary

Introduction

Oviposition sites[6,7,8,9,10], genes that code for proteins involved in olfactory reception may prove useful in this regard as well. The goal was to assess by qPCR the expression profile of three OBPs genes, Obp19d_2, Obp56a and Obp99c, at different developmental stages on the West Indies fruit fly. These genes were chosen due to their differential expression between virgin and post-mating males in a previous in silico analysis, suggesting them as potential targets for further studies. In addition to an analysis of OBPs’ expression, this study aimed to determine a set of reliable reference genes for expression studies across different developmental stages for the West Indies fruit fly. Our results show that some OBP genes are differentially expressed across different genders and reproductive stages, which make them good candidates for future application in genetic control

Objectives

Methods

Results

Conclusion