Gene expression of Mal d 1 allergen isoforms in apple fruits determined using new generation sequencing

Apples are one of the most consumed fruit species. However, eating fresh apples can cause an allergic reaction in a certain percentage of population. We evaluated the gene expression of Mal d 1, a major apple allergen in Central and Northern Europe, in four cultivars: ‘Idared’, UEB 32642, ‘Topaz’, and ‘Rubín’. Fruits of ‘Idared’, ‘Topaz’ and ‘Rubín’, were freshly harvested, while fruits of UEB 32642 were stored for four months in two ways: (i) under ultra-low oxygen conditions and (ii) under cold storage conditions. Total RNA was isolated from the apples and transcribed into cDNA. We applied the new generation sequencing (NGS) method using the Ion Torrent Personal Genome Machine (Ion PGM) sequencer to quantify the gene expression of all 31 allergen isoforms. Out of the tested allergen isoforms, relative gene expression exceeding 1% was observed only for seven genes, namely Mal d 1.01, Mal d 1.02, Mal d 1.05, Mal d 1.06A, Mal d 1.06B, Mal d 1.13A, and Mal d 1.13C. The expression of these genes accounted for 95% of all allergens contained at varying rates in the apples. However, only three genes, Mal d 1.01, Mal d 1.02 and Mal d 1.06A, dominated in all cultivars and represented 88-98% of all allergens of the cultivar. In contrast, almost no expression at background level was observed for three genes Mal d 1.04, Mal d 1.12 and Mal d 1.14.