Chlorate resistance analysis is an effective approach commonly used to distinguish the genetic variation between Oryza sativa L. ssp. indica and japonica, and predict the nitrogen use efficiency (NUE). This study aimed at investigating the response of a doubled haploid (DH) population derived from anther culture of 93-11 × Milyang352 exposed to 0.1% potassium chlorate (KClO3) at the seedling stage. The results revealed that the parental rice lines 93-11 (indica) and Milyang352 (japonica) showed distinctive phenotypic responses. The parental line 93-11 scored highly sensitive (0% survival) and Milyang352 scored resistant (66.7% survival) 7 days after treatment. The DH lines reflected the differential phenotypic response observed in parental lines. Interestingly, we identified a novel quantitative trait locus (QTL) for chlorate resistance on chromosome 3 (qCHR-3, 136 cM, logarithm of the odds—LOD: 4.1) using Kompetitive Allele-Specific PCR (KASP) markers. The additive effect (−11.97) and phenotypic variation explained (PVE; 14.9%) indicated that the allele from Milyang352 explained the observed phenotypic variation. In addition, shoot growth showed a significant difference between parental lines, but not root growth. Moreover, in silico analysis identified candidate genes with diverse and interesting molecular and physiological functions. Therefore, this study suggested that the QTL qCHR-3 harbors promising candidate genes that could play a role in the regulation of nitrogen metabolism in rice.
Potassium chlorate (KClO3) has been widely used to evaluate the divergence in nitrogen use efficiency (NUE) between indica and japonica rice subspecies. This study investigated the transcriptional regulation of major genes involved in the NUE in rice treated with KClO3, which acts as an inhibitor of the reducing activity of nitrate reductase (NR) in higher plants. A set of two KClO3 sensitive nitrate reductase (NR) and two nitrate transporter (NRT) introgression rice lines (BC2F7), carrying the indica alleles of NR or NRT, derived from a cross between Saeilmi (japonica, P1) and Milyang23 (indica, P2), were exposed to KClO3 at the seedling stage. The phenotypic responses were recorded 7 days after treatment, and samples for gene expression, physiological, and biochemical analyses were collected at 0 h (control) and 3 h after KClO3 application. The results revealed that Saeilmi (P1, japonica) and Milyang23 (P2, indica) showed distinctive phenotypic responses. In addition, the expression of OsNR2 was differentially regulated between the roots, stem, and leaf tissues, and between introgression lines. When expressed in the roots, OsNR2 was downregulated in all introgression lines. However, in the stem and leaves, OsNR2 was upregulated in the NR introgression lines, but downregulation in the NRT introgression lines. In the same way, the expression patterns of OsNIA1 and OsNIA2 in the roots, stem, and leaves indicated a differential transcriptional regulation by KClO3, with OsNIA2 prevailing over OsNIA1 in the roots. Under the same conditions, the activity of NR was inhibited in the roots and differentially regulated in the stem and leaf tissues. Furthermore, the transcriptional divergence of OsAMT1.3 and OsAMT2.3, OsGLU1 and OsGLU2, between NR and NRT, coupled with the NR activity pattern in the roots, would indicate the prevalence of nitrate (NO3¯) transport over ammonium (NH4+) transport. Moreover, the induction of catalase (CAT) and polyphenol oxidase (PPO) enzyme activities in Saeilmi (P1, KClO3 resistant), and the decrease in Milyang23 (P2, KClO3 sensitive), coupled with the malondialdehyde (MDA) content, indicated the extent of the oxidative stress, and the induction of the adaptive response mechanism, tending to maintain a balanced reduction–oxidation state in response to KClO3. The changes in the chloroplast pigments and proline content propose these compounds as emerging biomarkers for assessing the overall plant health status. These results suggest that the inhibitory potential of KClO3 on the reduction activity of the nitrate reductase (NR), as well as that of the genes encoding the nitrate and ammonium transporters, and glutamate synthase are tissue-specific, which may differentially affect the transport and assimilation of nitrate or ammonium in rice.
Lab-on-a-chip technology is an emerging and convenient system to easily and quickly separate proteins of high molecular weight. The current study established a high-molecular-weight glutenin subunit (HMW-GS) identification system using Lab-on-a-chip for three, six, and three of the allelic variations at the Glu-A1, Glu-B1, and Glu-D1 loci, respectively, which are commonly used in wheat breeding programs. The molecular weight of 1Ax1 and 1Ax2* encoded by Glu-A1 locus were of 200 kDa and 192 kDa and positioned below 1Dx subunits. The HMW-GS encoded by Glu-B1 locus were electrophoresed in the following order below 1Ax1 and 1Ax2*: 1Bx13 ≥ 1Bx7 = 1Bx7OE > 1Bx17 > 1By16 > 1By8 = 1By18 > 1By9. 1Dx2 and Dx5 showed around 4-kDa difference in their molecular weights, with 1Dy10 and 1Dy12 having 11-kDa difference, and were clearly differentiated on Lab-on-a-chip. Additionally, some of the HMW-GS, including 1By8, 1By18, and 1Dy10, having different theoretical molecular weights showed similar electrophoretic mobility patterns on Lab-on-a-chip. The relative protein amount of 1Bx7OE was two-fold higher than that of 1Bx7 or 1Dx5 and, therefore, translated a significant increase in the protein amount in 1Bx7OE. Similarly, the relative protein amounts of 8 & 10 and 10 & 18 were higher than each subunit taken alone. Therefore, this study suggests the established HMW-GS identification system using Lab-on-a-chip as a reliable approach for evaluating HMW-GS for wheat breeding programs.
A speed-breeding system using photoperiod characteristics has recently been developed to reduce the entire growth period in wheat. In this study, the entire growth period of four Korean varieties was examined to investigate whether this speed-breeding system would be beneficial for our wheat breeding program. When four varieties were cultivated under a 22-hour light/2-hour dark cycle in a glasshouse, the number of days to heading of Jokyoung and Baekkang was 44 and 43, respectively, and the number for Keumgang and Joongmo2008 was more than 75. Around twelve seeds per plant were obtained from Jokyoung and Baekkang, and the seeds of these varieties completely germinated when harvested at 20 days after heading. These results suggest that this speed-breeding system can be a reliable method of reducing the growth period in Korean wheat breeding.
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