Mutations affecting seed coat composition, structure, or morphology would likely affect its function.
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Hormones and possibly flavonoid compounds synthesized in the seed coat may also play a fundamental role in regulating communication between the seed coat and the endosperm and embryo. The hilum is the specialized area of the seed coat where the seed attaches to the pod and where nutrients are transferred from the pod to the seed. Other roles contributed by the seed coat in legumes and other seed plants include production of phenolic compounds as flavonoids, anthocyanins, and proanthocyanidins that have functions in defense, water permeability, dormancy, and germination of the seed. After that time dessication and yellowing proceeds until approximately 70 DAP, depending on the variety and maturity group with mature seeds of fresh weight of approximately 200 mg and 10% moisture content.ĭuring this process the seed coat (testa), which envelopes the cotyledons, develops from the ovule after fertilization and is essential in transferring assimilates and nutrients that allow the cotyledons to accumulate large amounts of storage proteins and oils in the soybean seed. The endosperm begins to degenerate and the cotyledons and embryo continue to grow for another 45 days and up to a maximum fresh weight of approximately 400–500 mg for the green seed. The embryo develops to reach the cotyledon phase 12 days after pollination (DAP). Three to 5 days after a flower is fertilized, the embryo with a globular shape is embedded in an acellular endosperm surrounded by a vascularized integument that provides the nutrition to both the embryo and endosperm while transforming itself into the seed coat. Soybean seed development is a complex process that initiates soon after flower fertilization. We present transcriptome and phenotypic evidence that substantially higher expression of an ethylene-forming ACO gene likely shifts hormone balance and sets in motion downstream changes resulting in a smaller hilum phenotype and the cracks observed in the minute hilum ( mi) isoline as compared to its recurrent parent.
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These changes are postulated to determine the restricted hilum size and cracking phenotypes. Evidence of changes in expression of genes downstream of the ethylene pathway included those involved in auxin and gibberellin hormone action and extensive differences in expression of cell wall protein genes. Glyma.09G008400 annotated as encoding an ethylene forming enzyme, ACC oxidase ( ACO), was found to be highly overexpressed in the mi hilum region at 165 RPKMs (reads per kilobase per million mapped reads) compared to the standard line at just 0.03 RPKMs.
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The RNA sequence data obtained from the 12 cDNA libraries were subjected to padj value < 0.05 and at least two-fold expression differences to select with confidence genes differentially expressed in the hilum-containing tissue of the seed coat between the two lines. The transcriptomes of these samples from both the Clark isoline containing the mi allele (PI 547628, UC413, i i R t mi G), and its recurrent Clark 63 parent isoline (PI 548532, UC7, i i R T Mi g), which was used for six generations of backcrossing, were compared for differential expression of 88,648 Glyma models of the soybean genome Wm82.a2. RNAs were extracted from immature seed from an extended hilum region (i.e., the hilum and a small ring of tissue surrounding the hilum in which the cracks form) at three different developmental stages:10–25, 25–50 and 50–100 mg seed fresh weight in two independent replicates for each stage. To elucidate features of seed development, we investigated the transcriptome of a soybean isoline from the germplasm collection that contained an introgressed allele known as minute hilum ( mi) which confers a smaller hilum region where the seed attaches to the pod and also results in seed coat cracking surrounding the hilum region.