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Allele-specific marker development and selection efficiencies for both flavonoid 3'-hydroxylase and flavonoid 3',5'-hydroxylase genes in soybean subgenus soja.

ABSTRACT: Color is one of the phenotypic markers mostly used to study soybean (Glycine max L. Merr.) genetic, molecular and biochemical processes. Two P450-dependent mono-oxygenases, flavonoid 3'-hydroxylase (F3'H; EC1.14.3.21) and flavonoid 3',5'-hydroxylase (F3'5'H, EC1.14.13.88), both catalyzing the hydroxylation of the B-ring in flavonoids, play an important role in coloration. Previous studies showed that the T locus was a gene encoding F3'H and the W1 locus co-segregated with a gene encoding F3'5'H in soybean. These two genetic loci have identified to control seed coat, flower and pubescence colors. However, the allelic distributions of both F3'H and F3'5'H genes in soybean were unknown. In this study, three novel alleles were identified (two of four alleles for GmF3'H and one of three alleles for GmF3'5'H). A set of gene-tagged markers was developed and verified based on the sequence diversity of all seven alleles. Furthermore, the markers were used to analyze soybean accessions including 170 cultivated soybeans (G. max) from a mini core collection and 102 wild soybeans (G. soja). For both F3'H and F3'5'H, the marker selection efficiencies for pubescence color and flower color were determined. The results showed that one GmF3'H allele explained 92.2 % of the variation in tawny and two gmf3'h alleles explained 63.8 % of the variation in gray pubescence colors. In addition, two GmF3'5'H alleles and one gmF3'5'h allele explained 94.0 % of the variation in purple and 75.3 % in white flowers, respectively. By the combination of the two loci, seed coat color was determined. In total, 90.9 % of accessions possessing both the gmf3'h-b and gmf3'5'h alleles had yellow seed coats. Therefore, seed coat colors are controlled by more than two loci.

SUBMITTER: Guo Y

PROVIDER: S-EPMC3664743 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundGlycine soja is a wild relative of soybean that has purple flowers. No flower color variant of Glycine soja has been found in the natural habitat.ResultsB09121, an accession with light purple flowers, was discovered in southern Japan. Genetic analysis revealed that the gene responsible for the light purple flowers was allelic to the W1 locus encoding flavonoid 3'5'-hydroxylase (F3'5'H). The new allele was designated as w1-lp. The dominance relationship of the locus was W1 >w1-lp >w1. One F2 plant and four F3 plants with purple flowers were generated in the cross between B09121 and a Clark near-isogenic line with w1 allele. Flower petals of B09121 contained lower amounts of four major anthocyanins (malvidin 3,5-di-O-glucoside, petunidin 3,5-di-O-glucoside, delphinidin 3,5-di-O-glucoside and delphinidin 3-O-glucoside) common in purple flowers and contained small amounts of the 5'-unsubstituted versions of the above anthocyanins, peonidin 3,5-di-O-glucoside, cyanidin 3,5-di-O-glucoside and cyanidin 3-O-glucoside, suggesting that F3'5'H activity was reduced and flavonoid 3'-hydroxylase activity was increased. F3'5'H cDNAs were cloned from Clark and B09121 by RT-PCR. The cDNA of B09121 had a unique base substitution resulting in the substitution of valine with methionine at amino acid position 210. The base substitution was ascertained by dCAPS analysis. The polymorphism associated with the dCAPS markers co-segregated with flower color in the F2 population. F3 progeny test, and dCAPS and indel analyses suggested that the plants with purple flowers might be due to intragenic recombination and that the 65 bp insertion responsible for gene dysfunction might have been eliminated in such plants.ConclusionsB09121 may be the first example of a flower color variant found in nature. The light purple flower was controlled by a new allele of the W1 locus encoding F3'5'H. The flower petals contained unique anthocyanins not found in soybean and G. soja. B09121 may be a useful tool for studies of the structural and functional properties of F3'5'H genes as well as investigations on the role of flower color in relation to adaptation of G. soja to natural habitats.

Project description:Background:Anthocyanin compounds playing multiple biological functions can be synthesized in different parts of barley (Hordeum vulgare L.) plant. The diversity of anthocyanin molecules is related with branching the pathway to alternative ways in which dihydroflavonols may be modified either with the help of flavonoid 3'-hydroxylase (F3'H) or flavonoid 3',5'-hydroxylase (F3'5'H)-the cytochrome P450-dependent monooxygenases. The F3'H and F3'5'H gene families are among the least studied anthocyanin biosynthesis structural genes in barley. The aim of this study was to identify and characterise duplicated copies of the F3'H and F3'5'H genes in the barley genome. Results:Four copies of the F3'5'H gene (on chromosomes 4HL, 6HL, 6HS and 7HS) and two copies of the F3'H gene (on chromosomes 1HL and 6HS) were identified in barley genome. These copies have either one or two introns. Amino acid sequences analysis demonstrated the presence of the flavonoid hydroxylase-featured conserved motifs in all copies of the F3'H and F3'5'H genes with the exception of F3'5'H-3 carrying a loss-of-function mutation in a conservative cytochrome P450 domain. It was shown that the divergence between F3'H and F3'5'H genes occurred 129 million years ago (MYA) before the emergence of monocot and dicot plant species. The F3'H copy approximately occurred 80 MYA; the appearance of F3'5'H copies occurred 8, 36 and 91 MYA. qRT-PCR analysis revealed the tissue-specific activity for some copies of the studied genes. The F3'H-1 gene was transcribed in aleurone layer, lemma and pericarp (with an increased level in the coloured pericarp), whereas the F3'H-2 gene was expressed in stems only. The F3'5'H-1 gene was expressed only in the aleurone layer, and in a coloured aleurone its expression was 30-fold higher. The transcriptional activity of F3'5'H-2 was detected in different tissues with significantly higher level in uncoloured genotype in contrast to coloured ones. The F3'5'H-3 gene expressed neither in stems nor in aleurone layer, lemma and pericarp. The F3'5'H-4 gene copy was weakly expressed in all tissues analysed. Conclusion:F3'H and F3'5'H-coding genes involved in anthocyanin synthesis in H. vulgare were identified and characterised, from which the copies designated F3'H-1, F3'H-2, F3'5'H-1 and F3'5'H-2 demonstrated tissue-specific expression patterns. Information on these modulators of the anthocyanin biosynthesis pathway can be used in future for manipulation with synthesis of diverse anthocyanin compounds in different parts of barley plant. Finding both the copies with tissue-specific expression and a copy undergoing pseudogenization demonstrated rapid evolutionary events tightly related with functional specialization of the duplicated members of the cytochrome P450-dependent monooxygenases gene families.

Project description:BackgroundGlycine soja is a halophytic soybean native to saline soil in Yellow River Delta, China. Photosystem I (PSI) performance and the interaction between photosystem II (PSII) and PSI remain unclear in Glycine soja under salt stress. This study aimed to explore salt adaptability in Glycine soja in terms of photosystems coordination.ResultsPotted Glycine soja was exposed to 300 mM NaCl for 9 days with a cultivated soybean, Glycine max, as control. Under salt stress, the maximal photochemical efficiency of PSII (Fv/Fm) and PSI (△MR/MR0) were significantly decreased with the loss of PSI and PSII reaction center proteins in Glycine max, and greater PSI vulnerability was suggested by earlier decrease in △MR/MR0 than Fv/Fm and depressed PSI oxidation in modulated 820 nm reflection transients. Inversely, PSI stability was defined in Glycine soja, as △MR/MR0 and PSI reaction center protein abundance were not affected by salt stress. Consistently, chloroplast ultrastructure and leaf lipid peroxidation were not affected in Glycine soja under salt stress. Inhibition on electron flow at PSII acceptor side helped protect PSI by restricting electron flow to PSI and seemed as a positive response in Glycine soja due to its rapid recovery after salt stress. Reciprocally, PSI stability aided in preventing PSII photoinhibition, as the simulated feedback inhibition by PSI inactivation induced great decrease in Fv/Fm under salt stress. In contrast, PSI inactivation elevated PSII excitation pressure through inhibition on PSII acceptor side and accelerated PSII photoinhibition in Glycine max, according to the positive and negative correlation of △MR/MR0 with efficiency that an electron moves beyond primary quinone and PSII excitation pressure respectively.ConclusionTherefore, photosystems coordination depending on PSI stability and rapid response of PSII acceptor side contributed to defending salt-induced oxidative stress on photosynthetic apparatus in Glycine soja. Photosystems interaction should be considered as one of the salt adaptable mechanisms in this halophytic soybean.

Project description:BACKGROUND AND AIMS:Wild soybean (Glycine soja), a native species of East Asia, is the closest wild relative of the cultivated soybean (G. max) and supplies valuable genetic resources for cultivar breeding. Analyses of the genetic variation and population structure of wild soybean are fundamental for effective conservation studies and utilization of this valuable genetic resource. METHODS:In this study, 40 wild soybean populations from China were genotyped with 20 microsatellites to investigate the natural population structure and genetic diversity. These results were integrated with previous microsatellite analyses for 231 representative individuals from East Asia to investigate the genetic relationships of wild soybeans from China. KEY RESULTS:Analysis of molecular variance (AMOVA) revealed that 43·92 % of the molecular variance occurred within populations, although relatively low genetic diversity was detected for natural wild soybean populations. Most of the populations exhibited significant effects of a genetic bottleneck. Principal co-ordinate analysis, construction of a Neighbor-Joining tree and Bayesian clustering indicated two main genotypic clusters of wild soybean from China. The wild soybean populations, which are distributed in north-east and south China, separated by the Huang-Huai Valley, displayed similar genotypes, whereas those populations from the Huang-Huai Valley were different. CONCLUSIONS:The previously unknown population structure of the natural populations of wild soybean distributed throughout China was determined. Two evolutionarily significant units were defined and further analysed by combining genetic diversity and structure analyses from Chinese populations with representative samples from Eastern Asia. The study suggests that during the glacial period there may have been an expansion route between south-east and north-east China, via the temperate forests in the East China Sea Land Bridge, which resulted in similar genotypes of wild soybean populations from these regions. Genetic diversity and bottleneck analysis supports that both extensive collection of germplasm resources and habitat management strategies should be undertaken for effective conservation studies of these important wild soybean resources.

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Allele-specific marker development and selection efficiencies for both flavonoid 3'-hydroxylase and flavonoid 3',5'-hydroxylase genes in soybean subgenus soja.

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