Reference gene selection for gene expression analysis in melon infected by Fusarium oxysporum f.sp. melonis

Abstract

Eleven candidate reference genes were evaluated in stem tissues of two melon (Cucumis melo L) genotypes infected with Fusarium oxysporum f.sp. melonis (FOM) race 1, to investigate the most stable internal control gene to use in gene expression study in melon under biotic stress. The candidate reference genes, utilized in the qRT-PCR, were selected among the most common reference genes reported in melon literature as well as on melon sequences found in available databases. The expression stability of the reference genes selected, was tested by using the geNorm and NormFinder statistical algorithms. Irrespective of the plant response to FOM infection, there was substantial agreement in gene ranks of both software. The ribosomal protein L2 gene had the high transcript stability while the elongation factor, cyclophilin7 and 18S rRNA were the least stable. The best three reference genes, ribosomal protein L2, actin* and cyclophilin were used to normalize the expression data of the glutathione S-transferase in the resistant and susceptible plants. Differences in the expression data emerged for both genotypes and were more evident in the resistant one. The ribosomal protein L2 showed similar normalized expression values in both genotypes and thus it could be indicated as unique internal control for gene expression analysis in melon under FOM infection. This work represents the first in-depth study on reference genes validation in melon under biotic stress condition. The identification of such genes would improve the accuracy and reliability of gene expression studies and speed up candidate disease resistant gene analysis in melon.