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Background  Improvement of α-tocopherol content is an important breeding aim to increase the nutritional value of crops. Several efforts have been conducted to improve the α-tocopherol content in soybean [ Glycine max  (L.) Merr.] through transgenic technology by overexpressing genes related to α-tocopherol biosynthesis or through changes to crop management practices. Varieties with high α-tocopherol content have been identified in soybean germplasms. The heritability of this trait has been characterized in a cross between high α-tocopherol variety Keszthelyi Aproszemu Sarga (KAS) and low α-tocopherol variety Ichihime. In this study, the genetic mechanism of the high α-tocopherol content trait of KAS was elucidated.    Results  Through QTL analysis and fine mapping in populations from a cross between KAS and a Japanese variety Ichihime, we identified  γ-TMT3 , which encodes  γ -tocopherol methyltransferase, as a candidate gene responsible for high α-tocopherol concentration in KAS. Several nucleotide polymorphisms including two nonsynonymous mutations were found in the coding region of  γ-TMT3  between Ichihime and KAS, but none of which was responsible for the difference in α-tocopherol concentration. Therefore, we focused on transcriptional regulation of  γ-TMT3  in developing seeds and leaves. An F 5  line that was heterozygous for the region containing  γ-TMT3  was self-pollinated. From among the progeny, plants that were homozygous at the  γ-TMT3  locus were chosen for further evaluation. The expression level of  γ-TMT3  was higher both in developing seeds and leaves of plants homozygous for the  γ-TMT3  allele from KAS. The higher expression level was closely correlated with high α-tocopherol content in developing seeds. We generated transgenic Arabidopsis plants harboring GUS gene under the control of  γ-TMT3  promoter from KAS or Ichihime. The GUS activity assay showed that the activity of  γ-TMT3  promoter from KAS was higher than that of Ichihime.    Conclusions  The genetic variation in  γ-TMT3 , which plays a major role in determining α-tocopherol concentration, provides significant information about the regulation of tocopherol biosynthesis in soybean seeds. This knowledge will help breeding programs to develop new soybean varieties with high α-tocopherol content.

Genetic variation of γ-tocopherol methyltransferase gene contributes to elevated α-tocopherol content in soybean seeds