Identification of Rf genes in hexaploid wheat (Triticum aestivum L.) by next generation sequencing of products of Representational Difference Analysis (RDA-seq).

Plants of two non-restorer varieties of hexaploid winter wheat (Astoria, Grana) and two restorers ones (Patres and Primépi) were used to Representational Difference Analysis (cDNA-RDA) to identify effective Rf (fertility restorer) genes. Anthers at the microspore stage were used as testers while anthers excised at the stage of pollen tetrads were selected as driver. The tester:driver ratio in three subsequent rounds of subtractive hybridization increased from 1:50, and 1:400 to 1:200000. Differential products obtained in the third subtraction of the size range from 200 to 800 bp were sequenced.