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Rice production relies on large amounts of synthetic nitrogen fertilizer to meet crop nutritional demands. However, the current fertilizer use efficiency is poor, with an average of 30–50% of total applied nitrogen (N) recovered in grains. Consequently, optimizing N availability and N use efficiency (NUE) has become a major target for rice yield improvement, and looking for quantitative trait loci (QTL) associated with NUE can provide useful information for developing nitrogen efficient rice genotypes. The present study aims to identify QTLs associated with NUE and its two components, agronomic NUE (agNUE) and grain nitrogen increase rate (GIR). A recombinant inbred line population (RIL) derived from a cross between Nipponbare and OM052, consisting of 159 F3 individuals was developed and cultivated in low-N (8 kg N/0.67 ha, N8) and high-N (18 kg N/0.67 ha, N18) conditions. Specific length amplified fragment sequencing (SLAF) was used to genotype the RILs and construct a high-density linkage map spanning 1587.70 cM, consisting 2707 SLAF markers with an average interval of 0.59 cM between adjacent marker loci. QTL analysis confirmed four significant QTLs distributed among chromosome 1, 6, and 11, with the proportion of phenotypic variance explained by each QTL ranging from 2.96 to 11.11% and LOD scores from 1.67 to 3.75. Functional annotation of genes located within the QTL intervals revealed NUE putative candidate genes. Overall, the QTLs identified in this study contribute information that could be useful for NUE improvement in rice.

Resistant starch (RS) is beneficial for human health, and especially for diabetics. Considering the high cost and low productivity of the Jiangtangdao 1 rice variety with high RS content, breeding high RS rice varieties exhibiting high productivity is essential. A molecular marker-assisted selection strategy was applied to increase RS content in a three-line hybrid rice variety. The functional rice variety Jiangtangdao 1, which contains sbe3-rs (on chr2) that controls the RS content, was used as the high RS content donor parent. Subsequently, male sterile maintainer and restorer lines containing homozygous sbe3-rs were bred using molecular marker-assisted selection combined with traditional breeding methods. The male sterile line was crossed with the restorer lines to identify the optimal hybrid combination with a high RS content. We obtained four combinations for which the yields were >50% higher than those of the control Jiangtangdao 1. In addition, there was no significant difference in the RS content between the combinations and Jiangtangdao 1. The hybrid rice plants with high RS content exhibited favorable agronomic traits and therefore have broad prospects for commercial application.

The acetohydroxy acid synthase S627N mutation confers herbicide tolerance in rice, and the rice variety containing this mutation produces good yields. This variety is commercially viable at Shanghai and Jiangsu regions in China. Weedy rice is a type of rice that produces lower yields and poorer quality grains than cultivated rice. It plagues commercial rice fields in many countries. One strategy to control its proliferation is to develop rice varieties that are tolerant to specific herbicides. Acetohydroxy acid synthase (AHAS) mutations have been found to confer herbicide tolerance to rice. Here, we identified a single mutation (S627N) in AHAS from an indica rice variety that conferred tolerance against imidazolinone herbicides, including imazethapyr and imazamox. A japonica rice variety (JD164) was developed to obtain herbicide tolerance by introducing the mutated indica ahas gene. Imidazolinone application was sufficient to efficiently control weedy rice in the JD164 field. Although the imazethapyr treatment caused dwarfing in the JD164 plants, it did not significantly reduce yields. To determine whether the decrease of the ahas mRNA expression caused the dwarfism of JD164 after imazethapyr application, we detected the ahas mRNA level in plants. The abundance of the ahas mRNA in JD164 increased after imidazolinone application, thus excluding the mRNA expression level as a possible cause of dwarfism. Activity assays showed that the mutated AHAS was tolerant to imidazolinone but the catalytic efficiency of the mutated AHAS decreased in its presence. Moreover, the activity of the mutated AHAS decreased more in the presence of imazethapyr than in the presence of imazamox. We observed no difference in the AHAS secondary structures, but homology modeling suggested that the S627N mutation enabled the substrate to access the active site channel in AHAS, resulting in imidazolinone tolerance. Our work combined herbicides with a rice variety to control weedy rice and showed the mechanism of herbicide tolerance in this rice variety.

Foods rich in resistant starch can help prevent various diseases, including diabetes, colon cancers, diarrhea, and chronic renal and hepatic diseases. Variations in starch biosynthesis enzymes could contribute to the high content of resistant starch in some cultivars of rice (Oryza sativa L.). Our previously published work indicated that the sbe3-rs gene in the rice mutant line, 'Jiangtangdao1' was a putative allele of the rice starch branching enzyme gene SBEIIb (previously known as SBE3); sbe3-rs might control the biosynthesis of the high resistant starch content in the rice line. Biomolecular analysis showed that the activity of SBEs was significantly lower in soluble extracts of immature seeds harvested from 'Jiangtangdao1' 15 days after flowering than in the extracts of the wild-type rice line 'Huaqingdao'. We performed gene complementation assays by introducing the wild-type OsSBEIIb into the sbe3-rs mutant 'Jiangtangdao1'. The genetically complemented lines demonstrated restored seed-related traits. The structures of endosperm amylopectin and the morphological and physicochemical properties of the starch granules in the transformants recovered to wild-type levels. This study provides evidence that sbe3-rs is a novel allele of OsSBEIIb, responsible for biosynthesis of high resistant starch in 'Jiangtangdao1'.

In the present study, the transgenic rice “Xiushui 134” with cry1Ac1 was developed by Agrobacterium mediated transformation method. Confirmed by PCR, all the 12 independent transformants were cry1Ac1 -positive plants. Using semi-quantitative reverse transcription PCR (RT-PCR), the cry1Ac1 gene expression could be detected in all independent transformants. Using real time quantitative PCR (qPCR) analysis, we found that 3 out of 12 transformants harbored a single copy cry1Ac1 gene. In order to study the integration site, the flanking sequences of these three transformants were amplified by thermal asymmetric interlaced (TAIL)-PCR. The result showed that the T-DNA of one of them inserted into genes interspace on the rice chromosome 2, about 2.4 kb upstream from the Os02G0539500 (Ch.2: 20858661~20860159) gene, and did not disturb other rice functional genes. This trans- formant was used as primary material to self-cross and segregate, and finally we got the homozygous cry1Ac1 lines. The resistance to leaf folder and agronomic traits of the lines were tested in field at natural conditions. The results showed that the homozygous cry1Ac1 rice plants had strong resistance to leaf folder while main agronomic traits were almost unaffected. These results indicate that we have successfully developed homozygous cry1Ac1 lines with highly resistant to leaf folder but main agronomic traits almost the same as controls.

Foods high in resistant starch (RS) are beneficial to prevent various diseases including diabetes, colon cancers, diarrhea and chronic renal or hepatic diseases. Elevated RS in rice is important for public health since rice is a staple food for half of the world population. A japonica mutant 'Jiangtangdao 1' (RS = 11.67%) was crossed with an indica cultivar 'Miyang 23' (RS = 0.41%). The mutant sbe3-rs that explained 60.4% of RS variation was mapped between RM6611 and RM13366 on chromosome 2 (LOD = 36) using 178 F(2) plants genotyped with 106 genome-wide polymorphic SSR markers. Using 656 plants from four F(3:4) families, sbe3-rs was fine mapped to a 573.3 Kb region between InDel 2 and InDel 6 using one STS, five SSRs and seven InDel markers. SBE3 which codes for starch branching enzyme was identified as a candidate gene within the putative region. Nine pairs of primers covering 22 exons were designed to sequence genomic DNA of the wild type for SBE3 and the mutant for sbe3-rs comparatively. Sequence analysis identified a missense mutation site where Leu-599 of the wild was changed to Pro-599 of the mutant in the SBE3 coding region. Because the point mutation resulted in the loss of a restriction enzyme site, sbe3-rs was not digested by a CAPS marker for SpeI site while SBE3 was. Co-segregation of the digestion pattern with RS content among 178 F(2) plants further supported sbe3-rs responsible for RS in rice. As a result, the CAPS marker could be used in marker-assisted breeding to develop rice cultivars with elevated RS which is otherwise difficult to accurately assess in crops. Transgenic technology should be employed for a definitive conclusion of the sbe3-rs.