Characterizing the expression of translation elongation factor gene EF1α in pear (Pyrus) fruit: evaluation of EF1α as a housekeeping gene

Reference genes are a key factor for the sensitivity and reliability of quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis. The elongation factor gene EF1α encodes a highly conserved ubiquitous protein that functions in the binding of aminoacyl-tRNA to the ribosome during peptide synthesis in eukaryotes, which is proposed for qRT-PCR as an internal reference for its putative housekeeping gene character. However, information about the paralogous copies and expression diversification of EF1α has been neglected and ill defined. In this study, the members of pear (Pyrus) EF1α were explored by mining public data. The family- and/or species-specific members of EF1α in Rosaceae plants or other species revealed by phylogenetic analysis suggested that the specific gene expansion occurred after the family or species diversification. The expression analysis based on the high-throughput sequencing and sequence differences typing provided a high resolution to distinguish the expression among the members of pear EF1α. The EF1α members showed an obviously unstable expression in both pear and apple (Malus × domestica) fruits at different developmental stages, clustered by two conserved expression patterns in pear fruit. The complementary expression among certain pear EF1α members leads to the sum expression of these members having a higher level of expression stability among the fruit at different developmental stages, which can be used as an approach to optimize EF1α as the internal reference. The result can also provide insight into the expression characteristics of pear EF1α in other tissues of pear. In addition, it indicates that any two genes having complementary expression patterns can be used as a reference for qPCR analysis with the high stability of their mean expression value.