Department of Biological Production, Akita Prefectural University, Akita, Japan.
Plant Mol Biol. 2022 Mar;108(4-5):325-342. doi: 10.1007/s11103-021-01161-9. Epub 2021 Jul 21.
Introduction of higher SSIIa activity to mild-type isa1 mutant by crossing results in restoration of crystallinity, starch granule structure, and production of plump seeds. Isoamylase 1 (ISA1) removes improper α-1, 6 glycosidic branches of amylopectin generated by starch branching enzymes and is essential for the formation of proper amylopectin structure. Rice isa1 (sug-1) mutants in japonica cultivar with less-active starch synthase IIa (SSIIa) and low granule-bound SSI (GBSSI) expression display wrinkled seed phenotype by accumulating water-soluble phytoglycogen instead of insoluble amylopectin. Expression of active SSIIa in transgenic rice produced with a severe-type isa1 mutant accumulated some insoluble glucan with weak B-type crystallinity at the periphery of seeds but their seeds remained wrinkled. To see whether introduction of high levels of SSIIa and/or GBSSI can restore the grain filling of the mild-type sug-1 mutant (EM653), new rice lines (SS2a gbss1 isa1, ss2a GBSS1 isa1, and SS2a GBSS1 isa1) were generated by crossing japonica isa1 mutant (ss2a gbss1 isa1) with wild type indica rice (SS2a GBSS1 ISA1). The results showed that SS2a gbss1 isa1 and SS2a GBSS1 isa1 lines generated chalky plump seeds accumulating insoluble amylopectin-like glucans with an increase in DP 13-35, while ss2a GBSS1 isa1 generated wrinkly seeds and accumulated soluble glucans enriched with DP < 13. Scanning electron microscopic observation of cross-section of the seeds showed that SS2a gbss1 isa1 and SS2a GBSS1 isa1 produced wild type-like polygonal starch granules. These starches showed the A-type crystallinity comparable to the wild type, while the japonica isa1 mutant and the transgenic rice do not show any or little crystallinity, respectively. These results indicate that introduction of higher SSIIa activity can mostly complements the mild-type sug-1 phenotype.
通过杂交将较高的 SSIIa 活性引入到轻症 isa1 突变体中，导致结晶度、淀粉颗粒结构和饱满种子的产生得到恢复。异淀粉酶 1（ISA1）去除淀粉分支酶产生的支链淀粉中不适当的α-1,6 糖苷键，并对形成适当的支链淀粉结构至关重要。粳稻品种 sug-1（sug-1）突变体中，SSIIa 活性较低，GBSSI（颗粒结合型 SS）表达量低，表现为皱粒表型，原因是积累了水溶性植酸糖原而不是不溶性支链淀粉。利用严重型 isa1 突变体进行的转基因水稻表达活性 SSIIa 后，在种子的外围积累了一些具有较弱 B 型结晶度的不溶性葡聚糖，但种子仍然皱缩。为了观察引入高活性 SSIIa 和/或 GBSSI 是否可以恢复轻症 sug-1 突变体（EM653）的灌浆，通过与野生型籼稻（SS2a GBSS1 ISA1）杂交，生成了新的水稻品系（SS2a gbss1 isa1、ss2a GBSS1 isa1 和 SS2a GBSS1 isa1）。结果表明，SS2a gbss1 isa1 和 SS2a GBSS1 isa1 系生成了垩白饱满的种子，积累了 DP13-35 增加的不溶性支链淀粉样葡聚糖，而 ss2a GBSS1 isa1 生成了皱粒种子并积累了富含 DP<13 的可溶性葡聚糖。种子横切面的扫描电子显微镜观察表明，SS2a gbss1 isa1 和 SS2a GBSS1 isa1 产生了与野生型相似的多角形淀粉颗粒。这些淀粉表现出与野生型相当的 A 型结晶度，而粳稻 isa1 突变体和转基因水稻则分别没有或几乎没有结晶度。这些结果表明，引入更高的 SSIIa 活性可以在很大程度上弥补轻症 sug-1 表型。
