Genotypic Variation in Spatial Distribution of Fe in Rice Grains in Relation to Phytic Acid Content and Ferritin Gene Expression

doi:10.1016/j.rsci.2020.04.005

Abstract

Abstract:

Rice varieties having high Fe concentration in the endospermic region can be used as a good source for Fe deficit population. In this study, 303 Oryza sativa varieties and 1 Oryza rufipogon accession were assessed for spatial Fe accumulation in grains by Prussian blue staining method. Spatial ferritin protein distribution in grains was visualized by immunohistochemistry, and ferritin expression was assessed in selected rice varieties using semi-quantitative reverse transcription PCR. Three popular rice varieties, namely Sarjoo 52, Madhukar and Jalmagna, and the O. rufipogon variety showed Fe in all the regions of grains, and the highest Fe concentration was observed in the embryo region. Some high-yielding varieties like Swarna, Swarna Sub 1, CSR13 and NDRR359 had lower Fe concentration in the embryo region. The highest Fe concentration was detected in O. rufipogon (49.8 μg/g), followed by Sarjoo 52 (26.1 μg/g) and Madhukar (25.7 μg/g). Phytic acid concentration was the minimum in O. rufipogon (5.75 mg/g) followed by Sarjoo 52 (5.83 mg/g). Western blot and semi-quantitative reverse transcription PCR showed higher expression of ferritin gene in O. rufipogon, Sarjoo 52 and Madhukar. In conclusion, O. rufipogon and Sarjoo 52 had higher Fe concentration in the embryo regions as well as endosperm and aleurone layer, whereas the other varieties had lower Fe concentration in the endosperm. Sarjoo 52 could be used as a donor in the rice breeding program for the generation of new varieties with elevated grain Fe concentration.

Key words: bio-fortification, iron accumulation, ferritin, tissue-specific localization, iron deficit population

Mishra Anurag, Shamim Md., Wasim Siddiqui Md., Singh Akanksha, Srivastava Deepti, N. Singh K.. Genotypic Variation in Spatial Distribution of Fe in Rice Grains in Relation to Phytic Acid Content and Ferritin Gene Expression[J]. Rice Science, 2020, 27(3): 227-236.

Figures/Tables 6

Fig. 1. Iron distribution and intensity in unpolished rice (A) and transverse section of matured rice grains (B) staining by Prussian blue method.i, Oryza rufipogon; ii, Sarjoo 52; iii, Swarna; iv, Swarna Sub 1; Em, Embryo; En, Endosperm; Al, Aleurone layer. Most of Fe (III) reaction is located in embryo part of the grains.

Fig. 2. Heat map on the basis of iron accumulation in different parts of rice grains.

Fig. 3. Correlation of iron and phytic acid contents in the rice genotypes.

Fig. 4. Immunolocalization of ferritin in different rice grains. A, Longitudinal section of Oryza rufipogon. B, Transverse section of Oryza rufipogon. C, Longitudinal section of Sarjoo 52. D, Transverse section of Sarjoo 52. E, Longitudinal section of Swarna. F, Transverse section of Swarna. G, Longitudinal section of Swarna Sub 1. H, Transverse section of Swarna Sub 1.Seed section imprints on nitrocellulose were visualized using ferritin antibody, accumulation and distribution.

Fig. 5. Western blot developed by the polyclonal ferritin antibody of the selected unpolished rice grains.

Fig. 6. Semi-quantitative RT-PCR analysis of different rice varieties.A, Ferritin expression in grains. B, Rice actin gene was used as template loading control on 1.5% agarose gel.1 to 12, O. rufipogon mature, O. rufipogon premature, Madhukar mature, Sarjoo 52 premature, Jalmagna mature, Jalmagna premature, Swarna mature, Swarna Sub 1 mature and Swarna Sub 1 premature grains, respectively.

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