N6 medium, Murashige and Skoog (MS) medium and BNBN medium were adopted in tissue cultures of Nipponbare (japonica) scutellum for performance comparison. Results indicated that, among the three kinds of culture media, the NB medium worked best in callus induction and gene transformation and was most suitable for tissue culture of Nipponbare scutellum. Based on this, the elicitor-encoding gene pemG1 from Magnaporthe grisea was introduced into the genome of Nipponbare via the Agrobacterium-mediated method. Transgenic rice plants were obtained. The integration, transcription and expression of pemG1 in rice were confirmed by polymerase chain reaction (PCR), Northern blot and Western blot, respectively. Genetic analysis showed that the transgenes were segregated normally in the progenies.

The development of (1→3,1→4)-β-glucan 4-glucanohydrolase (EC 3.2.1.73) has been examined in germinating wheat (Triticum aestivum cv. Millewa) grain, and in isolated aleurone layers and scutella. Activity is first detected in extracts of intact grain 2–3 days after the initiation of germination and thereafter increases until 6 days. In isolated aleurone layers, (1→3,1→4)-β-glucanase activity is secreted for up to 4 days. Treatment of aleurone layers with gibberellic acid (GA3) results in a 2-fold enhancement of secreted enzyme for the first 2 days, but activity decreases after 2 days. (1→3,1→4)-β-Glucanase secretion from GA3-treated aleurone layers clearly precedes the secretion of amylase and (1→4)-β-xylanase. Small but significant levels of cellulase are secreted from aleurone layers after GA3 treatment. Isolated scutella also secrete (1→3,1→4)-β-glucanase but GA3 has little apparent effect on secretion patterns from this tissue. Proteins secreted from excised aleurone layers and scutella, together with those in homogenates of intact, germinated grain, have been examined by Western blot analysis, using monoclonal antibodies against barley (1→3,1→4)-β-glucanases as probes. In each case the wheat (1→3,1→4)-β-glucanase is recognised only by the monoclonal antibody that is specific for barley isoenzyme El.

The rice (Oryza sativa L.) cv. Taichung 65, a japonica subspecies, was used to characterize the isoform, protein and gene expression patterns of endo-β-mannanase during and after seed germination. Activity assays and isoform analyses of whole grains or seed parts (scutellum, aleurone layer and starchy endosperm) revealed that seeds began to express endo-β-mannanase activity at 48 h from the start of imbibition at 25°C, after the completion of germination of most seeds. Three isoforms of endo-β-mannanase (pI 8.86, pI 8.92 and pI 8.98) were detected in the aleurone layer and starchy endosperm, but only two (pI 8.86 and pI 8.92) were present in the scutellum. The endo-β-mannanase in the starchy endosperm was mainly from the aleurone layer. Western blot analysis, using a tomato anti-endo-β-mannanase antibody, indicated that an endo-β-mannanase protein was present in an inactive form in dry grains. The amount of this protein decreased in the scutellum, but increased in the aleurone layer during and after germination. Thus, the increase in endo-β-mannanase activity in rice grains may be due to the activation of extant proteins and/or the de novo synthesis of the enzyme. Northern blot analysis showed that four putative rice endo-β-mannanase genes (OsMAN1, OsMAN2, OsMAN6 and OsMANP) were expressed in germinating and germinated rice grains. However, OsMANP was not expressed in the scutellum. The amount of OsMAN6 mRNA decreased after the completion of germination and paralleled the decline in endo-β-mannanase protein. In the aleurone layer, the increase of OsMAN2, OsMAN6 and OsMANP mRNA was prior to the increase of endo-β-mannanase protein.

Cereal grain germination and early seedling growth involve the co-ordinated action of endosperm and embryo tissues to mobilize the storage reserves of the starchy endosperm. This mobilization is accomplished by hydrolases secreted from the aleurone and scutellar tissues. The breakdown products are then transported to the growing seedling by the scutellum. This resource-harvesting system is regulated at multiple levels. One well-defined aspect of control is brought about by the hormone gibberellin (GA). Gibberellin is released from the embryo upon imbibition and activates the aleurone cells. The secretory apparatus of the aleurone then proliferates, supporting increased hydrolase synthesis and secretion to degrade the starchy endosperm. The molecules that regulate this response to GA are now being increasingly characterized. Elements such as cGMP, calcium, calmodulin and protein kinases are well known as regulators in other eukaryotic cell types and are emerging as key control factors in the aleurone hormone response. However, superimposed upon this molecular regulatory system is another level of control, the structural pattern of tissues and stored macro- molecules that was laid down during grain development. It is the interaction of these structural motifs combined with the molecular regulatory mechanisms that ensure the appropriate timing and positioning of hydrolase production and endosperm reserve mobilization. This integrated control system ensures an extended release of nutrients to fuel early seedling growth.

Peptide transport plays a major role in the nitrogen nutrition of the cereal embryo during germination. During germination, enzymatic hydrolysis of storage proteins in the cereal grain endosperm forms a reservoir of small peptides and amino acids which are translocated across the scutellum to supply organic nitrogen to the growing embryo. Uptake of these solutes by the scutellum, a modified cotyledon which functions in nutrient transport, is mediated by carriers localized to the plasma membrane of the scutellar epithelium. To date the peptide transporter HvPTR1 is by far the best characterized example of a nutrient transporter involved in reserve mobilization during germination. Peptide transport in the barley scutellum has been relatively well-characterized biochemically, and in recent years the barley scutellar peptide transporter HvPTR1 has been cloned and characterized at the molecular level. Here, we review the physiological role and importance of peptide transport in germination, focusing on recent characterization of the barley peptide transporter HvPTR1. In barley, the uptake of small peptides by the scutellum appears to be mediated by a proton-coupled peptide transport system capable of handling peptides 2–4 amino acid residues in length.