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Sugar content is related to fruit sweetness, and the complex mechanisms underlying fruit sugar accumulation still remain elusive. Here, we report a peach PpTST1 gene encoding tonoplast sugar transporter that is located in the quantitative trait loci (QTL) interval on Chr5 controlling fruit sucrose content. One derived Cleaved Amplified Polymorphic Sequence (dCAPS) marker was developed based on a nonsynonymous G/T variant in the third exon of PpTST1. Genotyping of peach cultivars with the dCAPS marker revealed a significant difference in fruit sugar content among genotypes. PpTST1 is located in the tonoplast, and substitution of glutamine by histidine caused by the G/T variation has no impact on subcellular location. The expression profile of PpTST1 exhibited a consistency with the sugar accumulation pattern, and its transient silencing significantly inhibited sugar accumulation in peach fruits. All of these results demonstrated the role of PpTST1 in regulating sugar accumulation in peach fruit. In addition, cis-elements for binding of MYB and WRKY transcript factors were found in the promoter sequence of PpTST1, suggesting a gene regulatory network of fruit sugar accumulation. Our results are not only helpful for understanding the mechanisms underlying fruit sugar accumulation, but will also be useful for the genetic improvement of fruit sweetness in peach breeding programs.

To enhance the marker density in the “QTL-hotspot” region, harboring several QTLs for drought tolerance-related traits identified on linkage group 04 (CaLG04) in chickpea recombinant inbred line (RIL) mapping population ICC 4958 × ICC 1882, a genotyping-by-sequencing approach was adopted. In total, 6.24 Gb data from ICC 4958, 5.65 Gb data from ICC 1882 and 59.03 Gb data from RILs were generated, which identified 828 novel single-nucleotide polymorphisms (SNPs) for genetic mapping. Together with these new markers, a high-density intra-specific genetic map was developed that comprised 1,007 marker loci spanning a distance of 727.29 cM. QTL analysis using the extended genetic map along with precise phenotyping data for 20 traits collected over one to seven seasons identified 49 SNP markers in the “QTL-hotspot” region. These efforts have refined the “QTL-hotspot” region to 14 cM. In total, 164 main-effect QTLs including 24 novel QTLs were identified. In addition, 49 SNPs integrated in the “QTL-hotspot” region were converted into cleaved amplified polymorphic sequence (CAPS) and derived CAPS (dCAPS) markers which can be used in marker-assisted breeding.

ABSTRACT Japanese brome (Bromus japonicus Houtt.) is a troublesome annual weed and widely distributed in winter wheat (Triticum aestivum L.) fields in the North China Plain. A B. japonicus population (TJ06) suspected of resistance to acetolactate synthase (ALS) inhibitors was found in Tianjin, China. In this study, the TJ06 population with an Asp-376-Glu mutation in ALS gene was identified. TJ06 population developed 66.7-fold resistance to flucarbazone-sodium and exhibited obvious cross-resistance to other two ALS-inhibiting herbicides. The 50% plant growth reduction (GR50) to herbicides of pyroxsulam and mesosulfuron-methyl were 28.72 and 39.44 g ai ha-1, respectively. In in vitro ALS activity assays, the concentration of flucarbazone-sodium required to inhibit 50% ALS activity (I50) for TJ06 was 11.3-fold greater than that for a susceptible population (TJ01), which was highly correlated with that of whole-plant response experiments and indicated that the Asp-376-Glu mutation leading to resistance reduced sensitivity of the ALS enzyme to flucarbazone- sodium. Besides, one derived cleaved amplified polymorphic sequence (dCAPS) marker was designed to quickly detect Asp376 mutation in ALS gene of B. japonicus.

High ploids of the sugarcane nuclear genome limit its genomic studies, whereas its chloroplast genome is small and conserved, which is suitable for phylogenetic studies and molecular marker development. Here, we applied whole genome sequencing technology to sequence and assemble chloroplast genomes of eight species of the ‘Saccharum Complex’, and elucidated their sequence variations. In total, 19 accessions were sequenced, and 23 chloroplast genomes were assembled, including 6 species of Saccharum (among them, S. robustum, S. sinense, and S. barberi firstly reported in this study) and 2 sugarcane relative species, Tripidium arundinaceum and Narenga porphyrocoma. The plastid phylogenetic signal demonstrated that S. officinarum and S. robustum shared a common ancestor, and that the cytoplasmic origins of S. sinense and S. barberi were much more ancient than the S. offcinarum/S. robustum linage. Overall, 14 markers were developed, including 9 InDel markers for distinguishing Saccharum from its relative species, 4 dCAPS markers for distinguishing S. officinarum from S. robustum, and 1 dCAPS marker for distinguishing S. sinense and S. barberi from other species. The results obtained from our studies will contribute to the understanding of the classification and plastome evolution of Saccharinae, and the molecular markers developed have demonstrated their highly discriminatory power in Saccharum and relative species.

The TGW6 gene encodes a novel indole-3-acetic acid-glucose hydrolase and plays a significant role in improving thousand grain weight (TGW) and yield in rice. In the current study, a TGW6 ortholog from bread wheat chromosome 3AL was isolated and designated as TaTGW6 - A1 . Sequencing and alignment analysis showed that the cloned TaTGW6 - A1 consists of a 987-bp intronless open reading frame. Three single-nucleotide polymorphisms (SNPs) were identified between cultivars with higher and lower TGW, forming two haplotypes, TaTGW6 - A1a and TaTGW6 - A1b . A derived cleaved amplified polymorphic sequence marker, TaTGW6 - A1 -CAPS, was developed to differentiate the two haplotypes. Linkage analysis using the wheat 90K iSelect SNP array mapped TaTGW6 - A1 -CAPS at 18 cM from BobWhite_c47304_56 in a RIL population derived from Zhou 8425B/Chinese Spring. A QTL for yield at the TaTGW6 - A1 locus explained 17.4 % of the phenotypic variance in average yield over four environments. Association analysis on 242 Chinese and foreign cultivars indicated that TaTGW6 - A1 was significantly associated with TGW. The cumulative frequency of the TaTGW6 - A1a allele associated with higher TGW was approximately 80 %, indicating that the TaTGW6 - A1a allele was positively selected in wheat breeding. In conclusion, TaTGW6 - A1 in conjunction with its molecular marker may contribute as a valuable gene in improving yield potential in wheat.

Barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.] is a dominant weed species occurring in rice (Oryza sativa L.) fields across China. Metamifop, a common herbicide, is frequently applied to control E. crus-galli and other grassy weeds in rice fields. Herein, HS01, an E. crus-galli population suspected to be resistant (R) to metamifop, was collected from Hanshan County in Anhui Province, China. Whole-plant dose–response testing revealed that, compared with the susceptible (S) population FD03, HS01 had developed high-level resistance to metamifop with a resistance index (RI) of 11.76 and showed cross-resistance to cyhalofop-butyl (RI = 9.33), fenoxaprop-P-ethyl (RI = 5.80) and clethodim (RI = 3.24). Gene sequencing revealed a Cys-2088-Arg mutation in the ACCase 1,5 allele of all the R plants, while ACCase gene overexpression was not involved in the resistance. Molecular docking indicated that the less-negative binding energies might be the main reason for the resistance of HS01 to acetyl-CoA carboxylase (ACCase)-inhibiting herbicides. A derived cleaved amplified polymorphic sequence (dCAPS) method was developed for the rapid identification of the Cys-to-Arg mutation in the ACCase gene at codon position 2088 in E. crus-galli. Additionally, pretreatment with the cytochrome P450 inhibitor piperonyl butoxide or the glutathione S-transferase inhibitor 4-chloro-7-nitrobenzoxadiazole had no significant effects (P > 0.05) on the resistance of HS01 to metamifop. To our knowledge, this is the first report of a Cys-2088-Arg mutation in E. crus-galli ACCase that confers cross-resistance to ACCase-inhibiting herbicides.

Chinese sprangletop [Leptochloa chinensis (L.) Nees] is one grass weed severely affecting rice (Oryza sativa L.) growth in paddies in China. Cyhalofop-butyl is the main herbicide used to control grass weeds in Chinese paddy fields, especially for controlling L. chinensis; however, L. chinensis has evolved resistance to cyhalofop-butyl due to continuous and extensive application. To investigate cyhalofop-butyl resistance levels and mechanisms in L. chinensis in some of the Chinese rice areas, 66 field populations were collected and treated with cyhalofop-butyl. Of these tested populations, 10 showed a high level of resistance to cyhalofop-butyl; the 50% effective dose ranged within 108.4 to 1,443.5 g ai ha–1 with resistance index values of 9.1 to 121.8 when compared with the susceptible population. Acetyl-coenzyme A carboxylase genes (ACCase) of susceptible and all 10 resistant populations were amplified and sequenced. Among them, Ile-1781-Leu, Trp-2027-Cys, Trp-2027-Ser, and Ile-2041-Asn mutations were found in five resistant populations. No known resistance-related mutations were found in the other five resistant populations, indicating that resistance to cyhalofop-butyl in these populations was likely to be endowed by non–target site resistance mechanisms. Notably, the Ile-1781-Leu and Trp-2027-Cys substitutions have previously been reported, but this is the first report of Trp-2027-Ser and Ile-2041-Asn mutations in L. chinensis. Furthermore, three derived cleaved amplified polymorphic sequence methods were developed to rapidly detect these mutations in L. chinensis.

Wheat stripe rust, caused by f. sp. , is a globally prevalent, wind-borne fungal disease and remains one of the most destructive threats to wheat production. By combining BSA-seq and QTL mapping using the wheat 120K SNP array, we identified two genomic regions associated with stripe rust resistance. Among them, a QTL on the short arm of chromosome 3A, , was consistently detected across environments, explaining 20.10%-25.21% of the phenotypic variance and showing LOD of 2.57-3.58. was delimited to an interval flanked by dCAPS-78 and dCAPS-83. Additive-effect analysis showed that RILs pyramiding with increased stripe rust resistance by 77.20% relative to RILs lacking both and , indicating that the pyramiding strategy had a significant impact on stripe rust resistance and underscoring its importance for high-yielding cultivars with durable resistance. The candidate interval exhibited high collinearity. Among the 48 high-confidence genes annotated in this region, an integrated analysis of transcriptome data, functional annotation, and sequence variation suggested that and are key candidate genes underlying . Collectively, these findings provide a foundation for marker-assisted breeding and future cloning and functional characterization of stripe rust resistance genes, which may help accelerate the development of elite stripe rust-resistant wheat cultivars, thereby improving wheat resistance to this destructive pathogen.

Weedy rice (Oryza sativa f. spontanea Roshev.), a widespread and troublesome weed in rice (Oryza sativa L.) fields, is typically controlled using imazamox in imidazolinone-tolerant rice fields. However, suspected resistance to imazamox has emerged in weedy rice populations in Jiangsu Province, China. This study aimed to evaluate the degree of resistance and investigate the resistance mechanisms. A whole-plant bioassay was performed on 35 weedy rice populations, demonstrating that 26 populations developed resistance to imazamox. The effective dose values causing 50% inhibition of growth reduction (GR50) in resistant (R) populations ranged from 129.2 to 280.2 g ai ha−1, exceeding the recommended application rate of imazamox (120 g ai ha−1) in imazamox-tolerant rice fields. R populations displayed cross-resistance to other acetolactate synthase (ALS)-inhibiting herbicides, except for certain sulfonylurea herbicides. Sequencing of the ALS gene identified a Ser-653-Asn substitution in resistant populations. A novel derived cleaved amplified polymorphic sequence (dCAPS) method was developed for the rapid and efficient detection of the Ser-653-Asn mutation in O. sativa f. spontanea. In vitro ALS activity assays revealed that the imazamox concentration required to inhibit 50% (IC50) of ALS activity was 80.0- to 88.3-fold higher in R populations compared with a susceptible (S) population. After imazamox treatment, the ALS expression levels in both the S and R populations of weedy rice increased. Resistance was not reversed by cytochrome P450 oxidase system (CYP450) or glutathione S-transferase (GST) inhibitors, suggesting that metabolic resistance mechanisms were not involved. In conclusion, weedy rice developed a different resistance level to imazamox, and the Ser-653-Asn mutation in the target ALS was the main reason. To the best of our knowledge, this study is the first to reveal the mechanism of resistance to imazamox in weedy rice in China.