The T locus of soybean (Glycine max (L.) Merr.) controls pubescence and seed coat color and is presumed to encode flavonoid 3'-hydroxylase (F3'H). The dominant T and the recessive t allele of the locus produce brown and gray pubescence, respectively. PCR primers were constructed based on the sequence of a soybean EST clone homologous to the F3'H gene. A putative full-length cDNA, sf3'h1 was isolated by 3' and 5' RACE. Sequence analysis revealed that sf3'h1 consists of 1690 nucleotides encoding 513 amino acids. It had 68% and 66% homology with corresponding F3'H protein sequences of petunia and Arabidopsis, respectively. A conserved amino acid sequence of F3'H proteins, GGEK, was found in the deduced polypeptide. Sequence analysis of the gene from a pair of near-isogenic lines for T, To7B (TT, brown) and To7G (tt, gray) revealed that they differed by a single C deletion in the coding region of To7G. The deletion changed the subsequent reading frame resulting in a truncated polypeptide lacking the GGEK consensus sequence and the heme-binding domain. Genomic Southern analysis probed by sf3'h1 revealed restriction fragment length polymorphisms between cultivars with different pubescence color. Further, sf3'h1 was mapped at the same position with T locus on LG3(c2). PCR-RFLP analysis was performed to detect the single-base deletion. To7B and three cultivars with brown pubescence exhibited shorter fragments, while To7G and three cultivars with gray pubescence had longer fragments due to the single-base deletion. The PCR-RFLP marker co-segregated with genotypes at the T locus in a F2 population segregating for the T locus. The above results strongly suggest that sJ3'h1 represents the T gene of soybean responsible for pubescence color and that the single-base deletion may be responsible for gray pubescence color.
Seed coats of soybean crack under various stress conditions. Cracking of seed coats degrades the external appearance of soybean seeds and reduces their commercial value. Previous studies revealed that the T gene responsible for pubescence color, and the maturity genes, E1 and E5, had inhibitory effects on low-temperature induced seed coat cracking. The objective of this study was to evaluate the effects of the T gene and five maturity genes (E1 to E5) on the intensity of seed coat cracking induced by pod-removal treatments. Soybean cv. Harosoy (te1e2E3E4e5) and its near-isogenic lines for T and E1 to E5 loci were used in the experiment. Cracking was induced by removing the upper 50% of pods on the plant 40 d after anthesis. Frequency and degree of cracking were not different among the isolines in the control group. In contrast, there were significant differences among isolines subjected to the pod-removal treatment. Frequency and degree of cracking was low in Harosoy, Harosoy-E1, e3, and e4, and high in Harosoy-T and E2. The results suggest that genotypes at T and E2 loci were associated with severity of seed coat cracking induced by pod-removal. There was a positive correlation (r = 0.90**) between individual seed weight and frequency of cracking among isolines in the pod-removal treatment. Seed coat cracking was probably exacerbated in part by the genes that allow enlargement of individual seeds in response to pod-removal. The differences among isolines suggest that the mechanism of seed coat cracking induced by pod removal may differ from that induced by low temperature treatment.
Exposure of soybean [Glycine max (L.) Merr.] to chilling temperatures at flowering stage induces browning around the hilum of the seed coats. The brown pigmentation spoils the external appearance of soybean seeds and reduces their commercial value. Our previous studies revealed that pigmentation was controlled by a few major genes, and one of the genes is closely associated with a maturity gene. This study was conducted to further investigate inheritance of pigmentation using DNA markers. Fifty-eight F(2) plants derived from a cross between a tolerant cv. Koganejiro and a sensitive cv. Kitakomachi were exposed to 15 degrees C for 2 weeks beginning 8 days after anthesis. Genotypes of 522 genetic markers were determined using the F(2) plants. Composite interval mapping revealed 5 quantitative trait loci (QTLs) for pigmentation, pig1 to pig5 (pig1 in molecular linkage group A2 [MLG A2], pig2 in MLG B1, pig3 in MLG C2, pig4 in molecular linkage group (MLG), and pig5 in MLG N) and 4 QTLs for flowering date, fd1 to fd4 (fd1 in MLG C1, fd2 in MLG C2, fd3 in MLG J, and fd4 in MLG L). Based on the relative location with markers, fd2 and fd4 probably correspond to E1 and E3, respectively. pig3 and fd2 were found at a similar position, and logarithm of odds (LOD) score plots for pigmentation and flowering date almost overlapped around this region. Considering the fact that pig3 had the most intense effects on pigmentation, E1 is presumed to be the maturity gene that profoundly affects pigmentation. Further, E3 has a small effect on pigmentation in accordance with the previous reports. These results support the idea that soybean maturity genes control low temperature-induced pigmentation with various intensities specific to each maturity gene. QTLs for seed coat pigmentation with small or no impact on maturity identified in this study may be useful in breeding for chilling tolerance.
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