Study On The Relationship Between SiPSY1 And Carotenoid Accumulation In Grains Of Foxtail Millet

The hulled grains of foxtail millet are referred to as millet,which is rich in nutrition.Carotenoids,as a nutrient of millet,have a great impact on millet color.Its more contents always responsible for the the yellower millet color and higher quality of millet.Phytoene synthase is the first key limiting enzyme in the carotenoid synthesis pathway.The expression patterns of SiPSY proteins and SiPSYs gene family members were studied in grains of different millet color varieties,and the relationship between the haplotypes of SiPSYs and the millet color of 343 Beijing resources were analyzed to find the gene which play the key role in carotenoid accumulation.This study will lay a theoretical foundation for the cultivating of high-quality millet varieties by molecular breeding.In this study,two white-millet varieties(ZSG and NMB)and two yellow-millet varieties(JG21 and CN35)were selected as the experimental materials.At different grain maturation stages,the millet color and ultrastructure of millet grains were checked,and the total carotenoids with its components,ABA contents,SiPSY protein and SiPSYs gene expression pattern between two millet color varieties were compared and analyzed.Furthermore,the relationship between the haplotypes of the SiPSY genes and the millet color were analyzed in 343 accessions.The molecular mechanism of SiPSY1 gene involved in regulating carotenoid synthesis in millet was revealed in depth from the physiological,biochemical and molecular levels.The main results were as follows:The millet color at different maturation stages was determined by colorimeter,and the results showed that the yellowness value b* of yellow millet varieties were significantly higher than that of white millet varieties.Laser confocal and transmission electron microscopy were used to observe and analyze the ultrastructure of millet grain.It was found that the contents of plastid pigment in the grains of different varieties were significantly different.The relative contents of amyloplast and plastid globules were related to the difference of carotenoid content in millet grains.The contents of total carotenoids,lutein,zeaxanthin,and ABA in grains of different varieties at different maturation stages were detected and compared by HPLC method.It was found that the contents of lutein and zeaxanthin in yellow millet varieties were significantly higher than those in white millet varieties.The contents of total carotenoids,lutein and zeaxanthin in yellow millet varieties were gradually increased with grain maturing,while those in white millet varieties were gradually decreased and zeaxanthin could hardly be detected at the last matured stage.The contents of ABA in grains of yellow millet varieties were decreased gradually with grain maturing,whereas gradually increased in grains of white millet varieties.The expression pattern of SiPSY protein was analyzed by western-blot,and it was found that the SiPSY protein expression in the grains of yellow and white millet varieties both increased to different degrees with grain maturing.In the end filling stage,the SiPSY protein expression of JG21 was significantly higher than that of NMB variety,and the SiPSY protein expression of ZSG was the highest in the middle and late,and end filling stage.The results of correlation analysis showed that high expression of SiPSY protein promoted carotenoid synthesis in yellow millet variety,and its overexpression might be related to excessive accumulation of ABA in white millet variety.The expression patterns of SiPSYs gene family members in grains at different maturation stages were analyzed by q RT-PCR,and the results showed that the expression of SiPSY1 gene in yellow millet varieties were all significantly higher than that in white millet varieties during the whole process of grain maturation.The results of correlation analysis showed that the expression of SiPSY1 gene in S3-S5 stages were positively correlated with the content of carotenoids and its components.Therefore,the over-expression of SiPSY1 gene in yellow millet varieties maybe one of the key factors to promote the accumulation of carotenoids leading to yellow millet color formation.The association analysis between the millet color of 343 Beijing resources and the haplotypes of the SiPSYs gene family revealed that there were significant differences among the major haplotypes of the SiPSY1 gene.For Hap5 of SiPSY1,there were base mutations and deletions in exon,intron and 5’UTR region in varieties with lower value of millet color.Comparing with other haplotypes found in millet varieties with higher millet color value,only this haplotype had a base mutation in the 5’UTR region of SiPSY1.Therefore,it may be one of the reasons for the difference of millet color.