Semi-dwarfism is a main agronomic trait in crop breeding. In this study, we performed genome-wide association study (GWAS) and identified a new quantitative trait nucleotide (QTN) for rice shoot length. The peak QTN (C/T) was located in the first coding region of a group III WRKY transcription factor OsWRKY21 (LOC_Os01g60640). Interestingly, further haplotype analysis showed that C/T difference only existed in the indica group but not in the japonica group, resulting in significant differences in plant height among the different indica rice varieties. OsWRKY21 was expressed in embryo, radicle, shoots, leaves, and stems. Most notably, overexpressing OsWRKY21 resulted in the semi-dwarf phenotype, early heading date and short internodes compared to the wild type, while the knockout mutant plants by CRISPR/Cas9 technology yielded the opposite. The overexpressing lines exhibited the decreased length of the cells near sclerenchyma epidermis, accompanied with the lower levels of indole-3-acetic acid (IAA) and gibberellin 3 (GA3), but increased levels of the abscisic acid (ABA) and salicylic acid (SA) in the internodes at heading stage. Moreover, the semi-dwarf phenotype could be fully rescued by exogenous GA3 application at seedling stage. The RNA-seq and qRT-PCR analysis confirmed the differential expression levels of genes in development and the stress responses in rice, including GA metabolism (GA20ox2, GA2ox6, and YABY1) and cell wall biosynthesis (CesA4, 7, and 9) and regulation (MYB103L). These data suggest the essential role of OsWRKY21 in regulation of internode elongation and plant height in rice.

Cold stress, including freezing stress and chilling stress, is one of the major environmental factors that limit the growth and productivity of plants. As a temperate dicot model plant species, Arabidopsis develops a capability to freezing tolerance through cold acclimation. The past decades have witnessed a deep understanding of mechanisms underlying cold stress signal perception, transduction, and freezing tolerance in Arabidopsis. In contrast, a monocot cereal model plant species derived from tropical and subtropical origins, rice, is very sensitive to chilling stress and has evolved a different mechanism for chilling stress signaling and response. In this review, the authors summarized the recent progress in our understanding of cold stress response mechanisms, highlighted the convergent and divergent mechanisms between Arabidopsis and rice plasma membrane cold stress perceptions, calcium signaling, phospholipid signaling, MAPK cascade signaling, ROS signaling, and ICE-CBF regulatory network, as well as light-regulated signal transduction system. Genetic engineering approaches of developing freezing tolerant Arabidopsis and chilling tolerant rice were also reviewed. Finally, the future perspective of cold stress signaling and tolerance in rice was proposed.

Marker-assisted backcrossing of a null allele of the a-subunit of Soybean (Glycine max) b-conglycinin with a Chinese soybean cultivar (a). The development of improved lines Abstract The development of soybean varieties that lack the b-conglycinin a-subunit is an attractive goal because the b-conglycinin a-subunit negatively influences the nutrition and gelation of tofu and is a major allergen. To remove this undesirable allergen and simultaneously improve the seed nutritional value and food-processing quality, marker-assisted background selection (MABS) was used in backcross breeding to incorporate cgy-2, a null phenotype version of the gene encoding the b-conglycinin a-subunit, from the donor line 'RiB' into the genetic background of the Chinese cultivar 'Dongnong47' (DN47), a popular high-oil superfine seed soybean cultivar from Heilongjiang Province, China. In each F 2 (F 2 , BC n F 2 ) generation of the breeding programme, the offspring that carried the introgressed cgy-2 were identified by sodium dodecyl sulphatepolyacrylamide gel electrophoresis and rescreened by MABS using simple sequence repeat markers to accelerate recurrent parent genome recovery. Of the 49 advanced backcrossing breeding lines (ABLs), the three best lines, ABL1, ABL2 and ABL3, were selected from the BC 1 , BC 2 and BC 3 populations, respectively. The ABLs were evaluated for desirable agronomic characteristics, yield-related traits, amino acid composition, free amino acid composition and tofu-processing quality in the mature seeds. All of the ABLs lacked the a-subunit but grew and reproduced normally without deleterious effects on physiological processes such as seed development and germination. The free amino acid content of ABL1 was significantly higher than that of 'DN47', with arginine (Arg) being particularly enriched. Compared to the recurrent parent 'DN47', the total protein content of the three ABLs was higher, the amino acid composition of the seed proteins was markedly modified and the yield and hardness of the tofu that was made from the ABLs were significantly increased. MABS combined with stringent phenotypic selection in a backcross breeding programme is a feasible strategy for the genetic engineering of seed protein components to produce allergenic subunit-deficient variant alleles.Key words: allergen a-subunit nullimproved lines -Soybean (Glycine max) b-conglycininsimple sequence repeat Plant Breeding, 133, 638-648 (2014)

Crossing, backcrossing, and molecular marker-assisted background selection produced a soybean (Glycine max) near-isogenic line (cgy-2-NIL) containing the cgy-2 allele, which is responsible for the absence of the allergenic α-subunit of β-conglycinin. To identify α-null-related transcriptional changes, the gene expressions of cgy-2-NIL and its recurrent parent DN47 were compared using Illumina high-throughput RNA-sequencing of samples at 25, 35, 50, and 55 days after flowering (DAF). Seeds at 18 DAF served as the control. Comparison of the transcript profiles identified 3,543 differentially expressed genes (DEGs) between the two genotypes, with 2,193 genes downregulated and 1,350 genes upregulated. The largest numbers of DEGs were identified at 55 DAF. The DEGs identified at 25 DAF represented a unique pattern of GO category distributions. KEGG pathway analyses identified 541 altered metabolic pathways in cgy-2-NIL. At 18DAF, 12 DEGs were involved in arginine and proline metabolism. The cgy-2 allele in the homozygous form modified the expression of several Cupin allergen genes. The cgy-2 allele is an alteration of a functional allele that is closely related to soybean protein amino acid quality, and is useful for hypoallergenic soybean breeding programs that aim to improve seed protein quality.

Spontaneous and radiation-induced mutants of soybean, despite loss of abundant seed proteins, have been reported to grow and reproduce normally without any apparent physiological abnormalities. Here, we report the development and characterization of a soybean line (BSH-2) that lacks several abundant seed storage proteins. One-dimensional and high-resolution two-dimensional gel electrophoresis revealed the absence of the α′ and α subunits of β-conglycinin and G1, G2, G4, and G5 glycinin in the newly developed mutant line (BSH-2). Like our earlier developed soybean mutant line (BSH-3), the seeds of BSH-2 also accumulated high levels of free amino acids as compared with wild-type DN47 seeds. An examination of the germination rates revealed that both BSH-2 and BSH-3 had significantly lower germination rates compared with the parent line DN47. Two-dimensional gel electrophoresis analysis demonstrated that these mutants had slower rates of mobilization of seed storage proteins. The delayed mobilization of storage proteins in BSH-2 and BSH-3 seeds was also correlated with a delayed induction of proteolytic activity in the mutants when compared to DN47. Similarly, qRT-PCR analysis revealed distinct expression pattern of genes involved in proteolytic pathway in the mutants when compared to DN47. Transmission electron microscopy observation of soybean seeds at two germination stages revealed striking differences in the breakdown of protein storage vacuoles and lipid bodies in the mutants. Our study demonstrates that BSH-2 and BSH-3 are compromised in mobilization of storage reserves and the absence of abundant storage proteins may affect the seed germination efficiency and post-germinative seedling establishment.