Search

Introduction: Current eggplant variety breeding is still mainly based on conventional methods, and there remains a lack of effective molecular breeding systems for complex traits controlled by multiple genes, such as yield and quality. To accelerate the research progress of eggplant genetics and molecular breeding, it is necessary to implement a genome-based breeding strategy. Methods: Therefore, in this study, a SNP array containing 50K liquid-phase probes was designed on the basis of the resequencing data of 577 eggplants. Results: The developed 50K liquid-phase probes were used to perform targeted capture sequencing on 12 eggplant lines, and the efficiency of probe capture exceeded 99.25%. Principal component, phylogenetic, and population structure analyses divided the 577 eggplants into 7 subgroups, and statistical analysis was performed on the fruit shape and color of the materials in the different subgroups. Further analysis of the geographical distribution of 428 Chinese eggplant materials revealed that the geographical regions of different subgroups were similar. The 50K SNP liquid-phase array was used to perform bulked- segregant analysis combined with whole-genome resequencing (BSA-seq) of fruit shape in the F2 population, which consisted of 1435 lines constructed with E421 as the maternal parent and 145 as the paternal parent. The BSA-seq data were located in the 78444173−84449348 interval on chromosome 3, with a size of 6 Mb, which was narrowed to 712.6 kb through fine mapping. Further sequence alignment and expression analysis revealed SmIQD14 as a candidate gene controlling eggplant fruit shape. The 50K SNP liquid-phase array can be widely used in future eggplant molecular breeding research. [ABSTRACT FROM AUTHOR]

In the ongoing arms race between rice and Magnaporthe oryzae, the pathogen employs effectors to evade the immune response, while the host develops resistance genes to recognise these effectors and confer resistance. In this study, we identified a novel Pik allele, Pik‐W25, from wild rice WR25 through bulked‐segregant analysis, creating the Pik‐W25 NIL (Near‐isogenic Lines) named G9. Pik‐W25 conferred resistance to isolates expressing AvrPik‐C/D/E alleles. CRISPR‐Cas9 editing was used to generate transgenic lines with a loss of function in Pik‐W25‐1 and Pik‐W25‐2, resulting in loss of resistance in G9 to isolates expressing the three alleles, confirming that Pik‐W25‐induced immunity required both Pik‐W25‐1 and Pik‐W25‐2. Yeast two‐hybrid (Y2H) and split luciferase complementation assays showed interactions between Pik‐W25‐1 and the three alleles, while Pik‐W25‐2 could not interact with AvrPik‐C, ‐D, and ‐E alleles with Y2H assay, indicating Pik‐W25‐1 acts as an adaptor and Pik‐W25‐2 transduces the signal to trigger resistance. The Pik‐W25 NIL exhibited enhanced field resistance to leaf and panicle blast without significant changes in morphology or development compared to the parent variety CO39, suggesting its potential for resistance breeding. These findings advance our knowledge of rice blast resistance mechanisms and offer valuable resources for effective and sustainable control strategies. Summary statement: We employed BSA‐seq to identify a novel Pik allele, Pik‐W25, from the wild rice WR25. Pik‐W25 can recognise AvrPik‐C, ‐D, or ‐E alleles and trigger resistance. Notably, the Pik‐W25 NIL showed enhanced field resistance to leaf and panicle blast without significant morphological or developmental differences. These findings advance our knowledge of rice blast resistance mechanisms and offer valuable resources for effective and sustainable rice blast control strategies. [ABSTRACT FROM AUTHOR]

Cold stress has a significantly negative effect on the growth, development, and yield of rice. However, the genetic basis for the differences in the cold tolerance of Xian/indica and Geng/japonica rice seedlings is still largely unknown. In this study, an RIL population was generated by crossing of the cold-tolerant japonica variety Nipponbare and the cold-sensitive indica variety WD16343 for BSA-seq analysis, and a major cold tolerance QTL qCTS11 was identified on chromosome 11. This locus was narrowed to the 584 kb region through fine mapping. Sequence alignment and expression analysis identified the cloned gene COLD11 and a novel cold-related gene OsCTS11. According to the reported functional variation of COLD11, Nipponbare (TCG + 3GCG)×2 presented more GCG repeats in the 1st exon than WD16343 (TCG + 3GCG), partially explaining the difference in cold tolerance between the parents. OsCTS11, encoding a stress enhanced protein based on phylogenetic analysis, was strongly induced by cold stress and located in the chloroplast and the nucleus. oscts11-mutant lines generated via CRISPR/Cas9 system improved the cold tolerance of rice seedlings. Our study not only reveals novel genetic loci associated with cold tolerance, but also provides potentially valuable gene resources for the cultivation of cold-tolerant rice. [ABSTRACT FROM AUTHOR]

Breeding for dwarfing traits in sorghum is crucial. However, only three genes (Dw1–Dw3) that control plant height have been mapped. In this study, 634 sorghum cultivars were collected to investigate plant height and genotypes. Four were genotyped Dw1DW2Dw3 (wild type) but with different plant heights, and they were selected to construct two populations and map new dwarf genes. Bulked segregant analysis with whole-genome resequencing of the two populations identified the candidate gene in one same genomic region—on chromosome 7. Then, it was narrowed down to a region containing nine genes. Amino acid and DNA sequence analysis of the parent and offspring plants revealed that two novel allelic mutations in the Dw3 gene play a role in reducing the plant height—8R262 or 8R417, including 1 bp substitution and 2 bp deletions. Furthermore, we sequenced 19 cultivars that primarily exhibited a "one-dwarf" hybrid or wild-type and presumed another allelic mutation via the amino acid alignment of 8R019, 8R100, and 8R402, which was another one-base substitution. These results indicate that multiple types of allelic mutations in the Dw3 gene should be considered when identified or applied. [ABSTRACT FROM AUTHOR]

Fruit color is an important qualitative trait that greatly influences the marketability of peppers. Fruit color can be divided into two categories. Green fruit color denotes commercial maturity, whereas ripe fruit indicates physiological maturity. Herein, segregation populations were created using the 'D24' with pale green in the green fruit stage, orange in the mature fruit stage, and 'D47' with green in the green fruit stage and red in the mature fruit stage. BSA-seq and genetic linkage map analysis revealed green fruit color was linked to (gyqtl1.1) on Chr1 and (gyqtl10.1) on Chr10, while mature fruit color was linked to Chr6. Using functional annotation, sequence, and expression analysis, we speculate that an SNP mutation in the CapGLK2 gene at the gyqtl10.1 interval could initiate premature termination of translation, thus yielding green to pale green fruits in D47. Conversely, the orange color in mature D24 fruits is due to the Indel-mediated premature termination of translation of the CapCCS gene. Our research offers a theoretical foundation for choosing different varieties of pepper fruit based on their color. [ABSTRACT FROM AUTHOR]

Exchangeable aluminum (Al) ions released from acidic soils with pH < 5.5 inhibit root elongation of crops, ultimately leading to yield reduced. It is necessary to identify the quantitative trait locus (QTLs) and candidate genes that confer toxicity resistance to understand the mechanism and improve tolerance of rapeseed. In this study, an F2 segregating population was derived from a cross between Al-tolerance inbred line FDH188 (R178) and -sensitive inbred line FDH152 (S169), and the F2:3 were used as materials to map QTLs associated with the relative elongation of taproot (RET) under Al toxicity stress. Based on bulked segregant analysis sequencing (BSA-seq), three QTLs (qAT-A07-1, qAT-A07-2, and qAT-A09-1) were detected as significantly associated with RET, and 656 candidate genes were screened. By combined BSA and RNA-seq analysis, 55 candidate genes showed differentially expressed, including genes encoding ABC transporter G (ABCG), zinc finger protein, NAC, ethylene-responsive transcription factor (ERF), etc. These genes were probably positive factors in coping with Al toxicity stress in rapeseed. This study provides new insight into exploring the QTLs and candidate genes' response to Al toxicity stress by combined BSA-seq and RNA-seq and is helpful to further research on the mechanism of Al resistance in rapeseed. [ABSTRACT FROM AUTHOR]

Crispness stands as a pivotal criterion in assessing apple texture, widely cherished by consumers. Yet, owing to its multifaceted nature, crispness remains a formidable challenge in artificial enhancement efforts. To expedite the early and precise evaluation of apple crispness, this study centered on a hybrid population derived from 'Fuji' and 'Pink Lady' cultivars, showcasing segregating crispness traits. We conducted measurements of flesh water content, cellular anatomical morphology, and employed a texture analyzer to assess mechanical properties of the offspring flesh. Integrating these three dimensions, we conducted a comprehensive analysis of quantitative characteristics of apple crispness, juxtaposed with sensory evaluation. Utilizing BSA-seq technology, we scrutinized extreme phenotypic individuals, revealing QTL loci intricately linked to the aforementioned dimensions, and subsequently developed Key Allele-Specific PCR (KASP) markers. These markers underwent validation in hybrid populations of 'Hanfu' x 'Pink Lady' and 'Hanfu' x 'Honey Crisp'. Our findings underscored significant correlations between mechanical properties, water content, and cell size with crispness. Higher mechanical properties and water content, alongside smaller cell size, correlated with firmer flesh texture; moderate mechanical properties, and elevated water content and cell size, with crisper texture; whereas lower mechanical properties, water content, and cell size implied softer flesh.The study yielded KASP markers effectively reflecting flesh mechanical properties (SNP_24399345), water content (SNP_8667563), and cell size (SNP_15566229). Comprehensive analysis of these markers identified CC-CC-TT as an effective identifier of soft flesh individuals; while GG-TC-TT and GG-CC-TT combinations better represented individuals with harder flesh. The Crunchy subclass could be discerned by combinations of GG-TC-TC, GG-TC-CC, GG-TT-TC, and GG-TT-CC. These findings furnish effective molecular markers for the genetic enhancement of apple crispness, bearing significant implications for the cultivation of novel apple varieties. [ABSTRACT FROM AUTHOR]

Hollowness is a physiological disorder that frequently occurs during the growth and postharvest storage phases of fleshy radish roots, significantly diminishing quality, yield, and marketability. However, the molecular mechanism for hollowness remains elusive. To identify the QTLs and potential candidate genes for hollowness tolerance in radish, F2 and BC1 populations were constructed from hollowness-tolerant radish (C16) and hollowness-sensitive radish (C17) in the present study. Genetic analysis indicated that hollowness tolerance may be governed by two independent recessive genes. By employing bulked segregant analysis sequencing (BSA-seq), two significant candidate genomic intervals were pinpointed on chromosomes R04 (960 kb, 6.48–7.44 Mb) and R05 (600 kb, 31.44–32.04 Mb), which together harbor 107 annotated genes. Transcriptomic sequencing revealed that the downregulated differentially expressed genes (DEGs) were significantly enriched in biological processes related to cell death and the response to water stress, whereas the upregulated DEGs were significantly associated with the chitin catabolic process and the cell wall macromolecule metabolic process. A total of 46 intersecting genes were identified among these DEGs within the genomic intervals of interest. One gene with high expression (Rsa10025345) and two with low expression (Rsa10025320 and Rsa10018106) were detected in the tolerant variety C16. Furthermore, a SNP within Rsa10025320 resulting in an amino acid change (A188E) was characterized through sequence variation observed in both BSA-seq and RNA-seq data and further developed as a derived cleaved amplified polymorphic sequence (dCAPS) marker. Our study reveals potential target genes for tolerance to hollowness and paves the way for marker-assisted breeding of hollowness tolerance in red-skinned radishes. [ABSTRACT FROM AUTHOR]

Soybean cyst nematode disease represents the major soil-borne disease of soybean. Identifying disease-resistant genes in soybean has a substantial impact on breeding of disease-resistant crops and genetic improvement. The present work created the F2 population with the disease-resistant line H-10 and disease-susceptible line Chidou4. 30 respective F2disease-resistant and disease-susceptible individuals for forming two DNA pools for whole-genome re-sequencing were selected. As a result, a total of 11,522,230 single nucleotide polymorphism (SNPs) markers from these two parental lines and two mixed pools were obtained. Accordng to SNP-index based association analysis, there were altogether 741 genes out of 99% confidence interval, which were mainly enriched into regions of 38,524,128∼39,849,988 bp with a total length of 1.33 Mb contain 111 genes on chromosome 2, 27,821,012∼29,612,574 bp with a total length of 1.79 Mb contain 92 genes on chromosome 3, 308∼348,214 bp with a total of length 0.35 Mb contain 34 genes on chromosome 10, and 53,867,581∼58,017, 852 bp with a total length of 4.15 Mb contain 504 genes on chromosome 18. Bulk segregant analysis in F2 generations (BSA-seq) was correlated with a disease resistance interval containing 15 genes. Then, using bioinformatics analysis and differential expression analysis, five candidate genes were identified: Glyma.02G211400, Glyma.18G252800, Glyma.18G285800, Glyma.18G287400 and Glyma.18G298200. Our results provides a key basis for analyzing the soybean resistance mechanism against soybean cyst nematode and cloning soybean resistance genes. [ABSTRACT FROM AUTHOR]

Abstract Cold stress has a significantly negative effect on the growth, development, and yield of rice. However, the genetic basis for the differences in the cold tolerance of Xian/indica and Geng/japonica rice seedlings is still largely unknown. In this study, an RIL population was generated by crossing of the cold-tolerant japonica variety Nipponbare and the cold-sensitive indica variety WD16343 for BSA-seq analysis, and a major cold tolerance QTL qCTS11 was identified on chromosome 11. This locus was narrowed to the 584 kb region through fine mapping. Sequence alignment and expression analysis identified the cloned gene COLD11 and a novel cold-related gene OsCTS11. According to the reported functional variation of COLD11, Nipponbare (TCG + 3GCG)×2 presented more GCG repeats in the 1st exon than WD16343 (TCG + 3GCG), partially explaining the difference in cold tolerance between the parents. OsCTS11, encoding a stress enhanced protein based on phylogenetic analysis, was strongly induced by cold stress and located in the chloroplast and the nucleus. oscts11-mutant lines generated via CRISPR/Cas9 system improved the cold tolerance of rice seedlings. Our study not only reveals novel genetic loci associated with cold tolerance, but also provides potentially valuable gene resources for the cultivation of cold-tolerant rice.

Cotton breeding for the development of early-maturing varieties is an effective way to improve multiple cropping indexes and alleviate the conflict between grains and cotton in the cultivated fields in China. In the present study, we aimed to identify upland cotton quantitative trait loci (QTLs) and candidate genes related to early-maturity traits, including whole growth period (WGP), flowering timing (FT), node of the first fruiting branch (NFFB), height of the node of the first fruiting branch (HNFFB), and plant height (PH). An early-maturing variety, CCRI50, and a late-maturing variety, Guoxinmian 11, were crossed to obtain biparental populations. These populations were used to map QTLs for the early-maturity traits for two years (2020 and 2021). With BSA-seq analysis based on the data of population 2020, the candidate regions related to early maturity were found to be located on chromosome D03. We then developed 22 polymorphic insertions or deletions (InDel) markers to further narrow down the candidate regions, resulting in the detection of five and four QTLs in the 2020 and 2021 populations, respectively. According to the results of QTL mapping, two candidate regions (InDel_G286-InDel_G144 and InDel_G24-InDel_G43) were detected. In these regions, three genes (GH_D03G0451, GH_D03G0649, and GH_D03G1180) have non-synonymous mutations in their exons and one gene (GH_D03G0450) has SNP variations in the upstream sequence between CCRI50 and Guoxinmian 11. These four genes also showed dominant expression in the floral organs. The expression levels of GH_D03G0451, GH_D03G0649 and GH_D03G1180 were significantly higher in CCRI50 than in Guoxinmian 11 during the bud differentiation stages, while GH_D03G0450 showed the opposite trend. Further functional verification of GH_D03G0451 indicated that the GH_D03G0451-silenced plants showed a delay in the flowering time. The results suggest that these are the candidate genes for cotton early maturity, and they may be used for breeding early-maturity cotton varieties.

Abstract As a Brassica crop, Brassica napus typically has single flowers that contain four petals. The double-flower phenotype of rapeseed has been a desirable trait in China because of its potential commercial value in ornamental tourism. However, few double-flowered germplasms have been documented in B. napus, and knowledge of the underlying genes is limited. Here, B. napus D376 was characterized as a double-flowered strain that presented an average of 10.92 ± 1.40 petals and other normal floral organs. F1, F2 and BC1 populations were constructed by crossing D376 with a single-flowered line reciprocally. Genetic analysis revealed that the double-flower trait was a recessive trait controlled by multiple genes. To identify the key genes controlling the double-flower trait, bulk segregant analysis sequencing (BSA-seq) and RNA-seq analyses were conducted on F2 individual bulks with opposite extreme phenotypes. Through BSA-seq, one candidate interval was mapped at the region of chromosome C05: 14.56–16.17 Mb. GO and KEGG enrichment analyses revealed that the DEGs were significantly enriched in carbohydrate metabolic processes, notably starch and sucrose metabolism. Interestingly, five and thirty-six DEGs associated with floral development were significantly up- and down-regulated, respectively, in the double-flowered plants. A combined analysis of BSA-seq and RNA-seq data revealed that five genes were candidates associated with the double flower trait, and BnaC05.ERS2 was the most promising gene. These findings provide novel insights into the breeding of double-flowered varieties and lay a theoretical foundation for unveiling the molecular mechanisms of floral development in B. napus.

Abstract Background The cold tolerance of rice is closely related to its production and geographic distribution. The identification of cold tolerance-related genes is of important significance for developing cold-tolerant rice. Dongxiang wild rice (Oryza rufipogon Griff.) (DXWR) is well-adapted to the cold climate of northernmost-latitude habitats ever found in the world, and is one of the most valuable rice germplasms for cold tolerance improvement. Results Transcriptome analysis revealed genes differentially expressed between Xieqingzao B (XB; a cold sensitive variety) and 19H19 (derived from an interspecific cross between DXWR and XB) in the room temperature (RT), low temperature (LT), and recovery treatments. The results demonstrated that chloroplast genes might be involved in the regulation of cold tolerance in rice. A high-resolution SNP genetic map was constructed using 120 BC5F2 lines derived from a cross between 19H19 and XB based on the genotyping-by-sequencing (GBS) technique. Two quantitative trait loci (QTLs) for cold tolerance at the early seedling stage (CTS), qCTS12 and qCTS8, were detected. Moreover, a total of 112 candidate genes associated with cold tolerance were identified based on bulked segregant analysis sequencing (BSA-seq). These candidate genes were divided into eight functional categories, and the expression trend of candidate genes related to ‘oxidation-reduction process’ and ‘response to stress’ differed between XB and 19H19 in the RT, LT and recovery treatments. Among these candidate genes, the expression level of LOC_Os12g18729 in 19H19 (related to ‘response to stress’) decreased in the LT treatment but restored and enhanced during the recovery treatment whereas the expression level of LOC_Os12g18729 in XB declined during recovery treatment. Additionally, XB contained a 42-bp deletion in the third exon of LOC_Os12g18729, and the genotype of BC5F2 individuals with a survival percentage (SP) lower than 15% was consistent with that of XB. Weighted gene coexpression network analysis (WGCNA) and modular regulatory network learning with per gene information (MERLIN) algorithm revealed a gene interaction/coexpression network regulating cold tolerance in rice. In the network, differentially expressed genes (DEGs) related to ‘oxidation-reduction process’, ‘response to stress’ and ‘protein phosphorylation’ interacted with LOC_Os12g18729. Moreover, the knockout mutant of LOC_Os12g18729 decreased cold tolerance in early rice seedling stage signifcantly compared with that of wild type. Conclusions In general, study of the genetic basis of cold tolerance of rice is important for the development of cold-tolerant rice varieties. In the present study, QTL mapping, BSA-seq and RNA-seq were integrated to identify two CTS QTLs qCTS8 and qCTS12. Furthermore, qRT-PCR, genotype sequencing and knockout analysis indicated that LOC_Os12g18729 could be the candidate gene of qCTS12. These results are expected to further exploration of the genetic mechanism of CTS in rice and improve cold tolerance of cultivated rice by introducing the cold tolerant genes from DXWR through marker-assisted selection.

Plant height (PH) is a critical agronomic trait in Brassica napus, significantly impacting yield. Consequently, identifying genes associated with plant height is a pivotal objective in oilseed rape breeding. This study employed a combination of bulk segregant analysis sequencing (BSA-seq) and RNA sequencing (RNA-seq) for analysis. A novel quantitative trait locus (QTL), qPH_C02, was identified between 63,989,634 and 64,945,122 bp on chromosome C02, from which eight candidate genes were screened. The Gene Ontology (GO) analysis revealed enrichment in peroxisomes, while the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated enrichment in the oxidative phosphorylation (OP) pathway. It is hypothesized that the observed differences in plant height and silique length may be attributed to the regulation of peroxidase activity in the OP pathway, which in turn alters plant energy metabolism and controls nutrient uptake. Subsequently, we will further test this hypothesis. The results of this study will contribute to our understanding of the genetic basis for differences in plant height and provide a foundation for the selection and breeding of Brassica napus varieties with desired plant shapes. [ABSTRACT FROM AUTHOR]

Soybean is a primary source of plant-based oil and protein for human diets. Seed size and weight are important agronomic traits that significantly influence soybean yield. Despite their importance, the genetic mechanisms underlying soybean seed size and weight remain to be fully elucidated. In order to identify additional, major quantitative trait loci (QTL) associated with seed size and weight, we developed segregating populations by crossing a large-seeded soybean variety "Kebaliang" with a small-seeded soybean variety "SUZUMARU". We evaluated seed length, width, thickness, and hundred-seed weight across two generations, F4 and F5, in 2022 and 2023. Employing bulked segregate analysis with whole-genome resequencing (BSA-seq), we detected 18 QTLs in the F4 population and 12 QTLs in the F5 population. Notably, six QTLs showed high stability between the two generations, with five derived from two pleiotropic loci (qSS4-1 and qSS20-1) and one specific to seed width (qSW14-1). Further validation and refinement of these loci were carried out through linkage mapping using molecular markers in the F5 population. Additionally, we identified 18 candidate genes within these stable loci and analyzed their sequence variations and expression profiles. Together, our findings offered a foundational reference for further soybean seed size research and unveiled novel genetic loci and candidate genes that could be harnessed for the genetic enhancement of soybean production. [ABSTRACT FROM AUTHOR]

Yellow leaf mutations have been widely used to study the chloroplast structures, the pigment synthesis, the photosynthesis mechanisms and the chlorophyll biosynthesis pathways across various species. For this study, a spontaneous mutant with the yellow leaf color named 96-140YBM was employed to explore the primary genetic elements that lead to the variations in the leaf color of hot peppers. To identify the pathways and genes associated with yellow leaf phenotypes, we applied sequencing-based Bulked Segregant Analysis (BSA-Seq) combined with BSR-Seq. We identified 4167 differentially expressed genes (DEGs) in the mutant pool compared with the wild-type pool. The results indicated that DEGs were involved in zeatin biosynthesis, plant hormone signal transduction, signal transduction mechanisms, post-translational modification and protein turnover. A total of 437 candidates were identified by the BSA-Seq, while the BSR-Seq pinpointed four candidate regions in chromosomes 8 and 9, containing 222 candidate genes. Additionally, the combination of BSA-Seq and BSR-Seq showed that there were 113 overlapping candidate genes between the two methods, among which 8 common candidates have been previously reported to be related to the development of chloroplasts, the photomorphogenesis and chlorophyll formation of plant chloroplasts and chlorophyll biogenesis. qRT-PCR analysis of the 8 common candidates showed higher expression levels in the mutant pool compared with the wild-type pool. Among the overlapping candidates, the DEG analysis showed that the CaKAS2 and CaMPH2 genes were down-regulated in the mutant pool compared to the wild type, suggesting that these genes may be key contributors to the yellow leaf phenotype of 96-140YBM. This research will deepen our understanding of the genetic basis of leaf color formation and provide valuable information for the breeding of hot peppers with diverse leaf colors. [ABSTRACT FROM AUTHOR]

As a Brassica crop, Brassica napus typically has single flowers that contain four petals. The double-flower phenotype of rapeseed has been a desirable trait in China because of its potential commercial value in ornamental tourism. However, few double-flowered germplasms have been documented in B. napus, and knowledge of the underlying genes is limited. Here, B. napus D376 was characterized as a double-flowered strain that presented an average of 10.92 ± 1.40 petals and other normal floral organs. F1, F2 and BC1 populations were constructed by crossing D376 with a single-flowered line reciprocally. Genetic analysis revealed that the double-flower trait was a recessive trait controlled by multiple genes. To identify the key genes controlling the double-flower trait, bulk segregant analysis sequencing (BSA-seq) and RNA-seq analyses were conducted on F2 individual bulks with opposite extreme phenotypes. Through BSA-seq, one candidate interval was mapped at the region of chromosome C05: 14.56–16.17 Mb. GO and KEGG enrichment analyses revealed that the DEGs were significantly enriched in carbohydrate metabolic processes, notably starch and sucrose metabolism. Interestingly, five and thirty-six DEGs associated with floral development were significantly up- and down-regulated, respectively, in the double-flowered plants. A combined analysis of BSA-seq and RNA-seq data revealed that five genes were candidates associated with the double flower trait, and BnaC05.ERS2 was the most promising gene. These findings provide novel insights into the breeding of double-flowered varieties and lay a theoretical foundation for unveiling the molecular mechanisms of floral development in B. napus. [ABSTRACT FROM AUTHOR]

Rising temperature is a major threat to the normal growth and development of maize, resulting in low yield production and quality. The mechanism of maize in response to heat stress remains uncertain. In this study, a maize mutant Zmhsl-1 (heat sensitive leaves) with wilting and curling leaves under high temperatures was identified from maize Zheng 58 (Z58) mutant lines generated by ethyl methanesulfonate (EMS) mutagenesis. The Zmhsl-1 plants were more sensitive to increased temperature than Z58 in the field during growth season. The Zmhsl-1 plants had lower plant height, lower yield, and lower content of photosynthetic pigments. A bulked segregant analysis coupled with whole-genome sequencing (BSA-seq) enabled the identification of the corresponding gene, named ZmHSL, which encodes an endo-β-1,4-xylanase from the GH10 family. The loss-of-function of ZmHSL resulted in reduced lignin content in Zmhsl-1 plants, leading to defects in water transport and more severe leaf wilting with the increase in temperature. RNA-seq analysis revealed that the differentially expressed genes identified between Z58 and Zmhsl-1 plants are mainly related to heat stress-responsive genes and unfolded protein response genes. All these data indicated that ZmHSL plays a key role in lignin synthesis, and its defective mutation causes changes in the cell wall structure and gene expression patterns, which impedes water transport and confers higher sensitivity to high-temperature stress. [ABSTRACT FROM AUTHOR]

Plant height, as a crucial component of plant architecture, exerts a significant influence on rapeseed (Brassica napus L.) lodging resistance, photosynthetic efficiency, yield, and mechanized harvest level. A previous study identified dwarf rapeseed LSW2018. In this study, LSW2018 (dwarf parent (PD)) was crossed with 389 (high parent (PH)) to establish the F2 population, and 30 extremely dwarf (bulk–D) and high (bulk–H) plants in the F2 population were respectively selected to construct two bulked DNA pools. Whole–genome sequencing and variation analysis (BSA–seq) were performed on these four DNA pools (PD, PH, bulk–D, and bulk–H). The BSA–seq results revealed that the genomic region responsible for the dwarf trait spanned from 19.30 to 22.19 Mb on chromosome A03, with a length of 2.89 Mb. After fine mapping with simple sequence repeat (SSR) markers, the gene was narrowed to a 0.71 Mb interval. Within this region, a total of 113 genes were identified, 42 of which contained large–effect variants. According to reference genome annotation and qRT–PCR analysis, there are 17 differentially expressed genes in this region between high and dwarf individuals. This study preliminarily reveals the genetic basis of LSW2018 dwarfing and provides a theoretical foundation for the molecular marker–assisted breeding of dwarf rapeseed. [ABSTRACT FROM AUTHOR]

Blade length, which is a quantitative trait controlled by multiple genes, is a key trait determining the yield in Pyropia haitanensis. However, there are few studies on the identification of candidate genes associated with it. In this study, the double haploid (DH) strains from the hybrid offspring of P. haitanensis were used to construct two DNA pools from extremely long-length and short-length strains, respectively. Using bulk segregant analysis sequencing (BSA-seq), three candidate regions associated with the blade length were identified on the genome, all of which were located on chromosome 1. These regions contain a total of 625 genes. One extremely long-length strain and one extremely short-length strain were selected respectively, and their 35-day blades were used for RNA sequencing (RNA-seq). It was found that the genes related to carbon fixation in photosynthetic organisms, glycolysis/gluconeogenesis and carbon metabolism were up-regulated in long blades, and 145 genes were differentially expressed in the candidate regions. Subsequently, eight candidate genes were screened using strict criteria (delta SNP-index ≥ 0.9), which encode WD40-repeat containing protein, major facilitator superfamily, LPXTG-anchored surface-anchored adhesin, RmlC-like cupin superfamily, EMP24_GP25L and three unknown functional proteins, respectively. Further SNP molecular marker verification showed that the success rate of selection of these 8 genes for blade length was greater than 90%. In addition, among the differentially expressed genes within the candidate regions, the expression levels of growth-related genes (PKCI and ABC transporters) were significantly different between 5 extremely long-length and 5 extremely short-length strains. In summary, this study identified 10 potential candidate genes associated with the blade length of P. haitanensis, thereby providing valuable insights for future advancements in precise molecular breeding and expedited development of high-yield varieties. [ABSTRACT FROM AUTHOR]

Fruit size is a crucial agronomic trait in bottle gourd, impacting both yield and utility. Despite its significance, the regulatory mechanism governing fruit size in bottle gourd remains largely unknown. In this study, we used bottle gourd (small-fruited H28 and large-fruited H17) parent plants to measure the width and length of fruits at various developmental stages, revealing a single 'S' growth curve for fruit expansion. Paraffin section observations indicated that both cell number and size significantly influence bottle gourd fruit size. Through bulked segregant analysis and combined genotype–phenotype analysis, the candidate interval regulating fruit size was pinpointed to 17,747,353 bp–18,185,825 bp on chromosome 9, encompassing 0.44 Mb and including 44 genes. Parental fruits in the rapid expansion stage were subjected to RNA-seq, highlighting that differentially expressed genes were mainly enriched in pathways related to cell wall biosynthesis, sugar metabolism, and hormone signaling. Transcriptome and resequencing analysis, combined with gene function annotation, identified six genes within the localized region as potential regulators of fruit size. This study not only maps the candidate interval of genes influencing fruit size in bottle gourd through forward genetics, but also offers new insights into the potential molecular mechanisms underlying this trait through transcriptome analysis. [ABSTRACT FROM AUTHOR]

Background: The cold tolerance of rice is closely related to its production and geographic distribution. The identification of cold tolerance-related genes is of important significance for developing cold-tolerant rice. Dongxiang wild rice (Oryza rufipogon Griff.) (DXWR) is well-adapted to the cold climate of northernmost-latitude habitats ever found in the world, and is one of the most valuable rice germplasms for cold tolerance improvement. Results: Transcriptome analysis revealed genes differentially expressed between Xieqingzao B (XB; a cold sensitive variety) and 19H19 (derived from an interspecific cross between DXWR and XB) in the room temperature (RT), low temperature (LT), and recovery treatments. The results demonstrated that chloroplast genes might be involved in the regulation of cold tolerance in rice. A high-resolution SNP genetic map was constructed using 120 BC5F2 lines derived from a cross between 19H19 and XB based on the genotyping-by-sequencing (GBS) technique. Two quantitative trait loci (QTLs) for cold tolerance at the early seedling stage (CTS), qCTS12 and qCTS8, were detected. Moreover, a total of 112 candidate genes associated with cold tolerance were identified based on bulked segregant analysis sequencing (BSA-seq). These candidate genes were divided into eight functional categories, and the expression trend of candidate genes related to 'oxidation-reduction process' and 'response to stress' differed between XB and 19H19 in the RT, LT and recovery treatments. Among these candidate genes, the expression level of LOC_Os12g18729 in 19H19 (related to 'response to stress') decreased in the LT treatment but restored and enhanced during the recovery treatment whereas the expression level of LOC_Os12g18729 in XB declined during recovery treatment. Additionally, XB contained a 42-bp deletion in the third exon of LOC_Os12g18729, and the genotype of BC5F2 individuals with a survival percentage (SP) lower than 15% was consistent with that of XB. Weighted gene coexpression network analysis (WGCNA) and modular regulatory network learning with per gene information (MERLIN) algorithm revealed a gene interaction/coexpression network regulating cold tolerance in rice. In the network, differentially expressed genes (DEGs) related to 'oxidation-reduction process', 'response to stress' and 'protein phosphorylation' interacted with LOC_Os12g18729. Moreover, the knockout mutant of LOC_Os12g18729 decreased cold tolerance in early rice seedling stage signifcantly compared with that of wild type. Conclusions: In general, study of the genetic basis of cold tolerance of rice is important for the development of cold-tolerant rice varieties. In the present study, QTL mapping, BSA-seq and RNA-seq were integrated to identify two CTS QTLs qCTS8 and qCTS12. Furthermore, qRT-PCR, genotype sequencing and knockout analysis indicated that LOC_Os12g18729 could be the candidate gene of qCTS12. These results are expected to further exploration of the genetic mechanism of CTS in rice and improve cold tolerance of cultivated rice by introducing the cold tolerant genes from DXWR through marker-assisted selection. [ABSTRACT FROM AUTHOR]

IntroductionBrassica juncea is a major oilseed crop of Brassica. The seed weight is one of yield components in oilseed Brassica crops. Research on the genetic mechanism of seed weight is not only directly related to the yield and economic value of Brassica juncea but also can provide a theory foundation for studying other Brassica crops.MethodsTo map the genes for seed weight, the parental and F2 extreme bulks derived were constructed from the cross between the heavy-seeded accession 7981 and the light-seeded one Sichuan yellow (SY) of B. juncea, and used in bulk segregant sequencing (BSA-seq). Meanwhile, RNA-sequencing (RNA-seq) was performed for both parents at six seed development stages.ResultsOur results showed that a total of thirty five SNPs were identified in thirty two genes located on chromosomes A02 and A10, while fifty eight InDels in fifty one genes located on A01, A03, A05, A07, A09, A10, B01, B02 and B04. The 7,679 differentially expressed genes were identified in developing seeds between the parents. Furthermore, integrated analysis of BSA-seq and RNA-seq data revealed a cluster of nine genes on chromosome A10 and one gene on chromosome A05 that are putative candidate genes controlling seed weight in B. juncea.DiscussionThis study provides a new reference for research on Brassica seed weight and lays a solid foundation for the examination of seed in other Brassica crops.

IntroductionCurrent eggplant variety breeding is still mainly based on conventional methods, and there remains a lack of effective molecular breeding systems for complex traits controlled by multiple genes, such as yield and quality. To accelerate the research progress of eggplant genetics and molecular breeding, it is necessary to implement a genome-based breeding strategy.MethodsTherefore, in this study, a SNP array containing 50K liquid-phase probes was designed on the basis of the resequencing data of 577 eggplants.ResultsThe developed 50K liquid-phase probes were used to perform targeted capture sequencing on 12 eggplant lines, and the efficiency of probe capture exceeded 99.25%. Principal component, phylogenetic, and population structure analyses divided the 577 eggplants into 7 subgroups, and statistical analysis was performed on the fruit shape and color of the materials in the different subgroups. Further analysis of the geographical distribution of 428 Chinese eggplant materials revealed that the geographical regions of different subgroups were similar. The 50K SNP liquid-phase array was used to perform bulked- segregant analysis combined with whole-genome resequencing (BSA-seq) of fruit shape in the F2 population, which consisted of 1435 lines constructed with E421 as the maternal parent and 145 as the paternal parent. The BSA-seq data were located in the 78444173−84449348 interval on chromosome 3, with a size of 6 Mb, which was narrowed to 712.6 kb through fine mapping. Further sequence alignment and expression analysis revealed SmIQD14 as a candidate gene controlling eggplant fruit shape. The 50K SNP liquid-phase array can be widely used in future eggplant molecular breeding research.

Fruit color is an important qualitative trait that greatly influences the marketability of peppers. Fruit color can be divided into two categories. Green fruit color denotes commercial maturity, whereas ripe fruit indicates physiological maturity. Herein, segregation populations were created using the ‘D24’ with pale green in the green fruit stage, orange in the mature fruit stage, and ‘D47’ with green in the green fruit stage and red in the mature fruit stage. BSA-seq and genetic linkage map analysis revealed green fruit color was linked to (gyqtl1.1) on Chr1 and (gyqtl10.1) on Chr10, while mature fruit color was linked to Chr6. Using functional annotation, sequence, and expression analysis, we speculate that an SNP mutation in the CapGLK2 gene at the gyqtl10.1 interval could initiate premature termination of translation, thus yielding green to pale green fruits in D47. Conversely, the orange color in mature D24 fruits is due to the Indel-mediated premature termination of translation of the CapCCS gene. Our research offers a theoretical foundation for choosing different varieties of pepper fruit based on their color.

Soybean cyst nematode disease represents the major soil-borne disease of soybean. Identifying disease-resistant genes in soybean has a substantial impact on breeding of disease-resistant crops and genetic improvement. The present work created the F2 population with the disease-resistant line H-10 and disease-susceptible line Chidou4. 30 respective F2disease-resistant and disease-susceptible individuals for forming two DNA pools for whole-genome re-sequencing were selected. As a result, a total of 11,522,230 single nucleotide polymorphism (SNPs) markers from these two parental lines and two mixed pools were obtained. Accordng to SNP-index based association analysis, there were altogether 741 genes out of 99% confidence interval, which were mainly enriched into regions of 38,524,128∼39,849,988 bp with a total length of 1.33 Mb contain 111 genes on chromosome 2, 27,821,012∼29,612,574 bp with a total length of 1.79 Mb contain 92 genes on chromosome 3, 308∼348,214 bp with a total of length 0.35 Mb contain 34 genes on chromosome 10, and 53,867,581∼58,017, 852 bp with a total length of 4.15 Mb contain 504 genes on chromosome 18. Bulk segregant analysis in F2 generations (BSA-seq) was correlated with a disease resistance interval containing 15 genes. Then, using bioinformatics analysis and differential expression analysis, five candidate genes were identified: Glyma.02G211400, Glyma.18G252800, Glyma.18G285800, Glyma.18G287400 and Glyma.18G298200. Our results provides a key basis for analyzing the soybean resistance mechanism against soybean cyst nematode and cloning soybean resistance genes.

Abstract Globe artichoke (Cynara cardunculus var. scolymus; 2n = 2x = 34) is a food crop consumed for its immature flower heads. Traditionally, globe artichoke varietal types are vegetatively propagated. However, seed propagation makes it possible to treat the crop as annual, increasing field uniformity and reducing farmers costs, as well as pathogens diffusion. Despite globe artichoke’s significant agricultural value and the critical role of heterosis in the development of superior varieties, the production of hybrids remains challenging without a reliable system for large-scale industrial seed production. Male sterility (MS) presents a promising avenue for overcoming these challenges by simplifying the hybridization process and enabling cost-effective seed production. However, within the Cynara genus, genic male sterility has been linked to three recessive loci in globe artichoke, with no definitive genetic mechanism elucidated to date. A 250 offsprings F2 population, derived from a cross between a MS globe artichoke and a male fertile (MF) cultivated cardoon (C. cardunculus var. altilis) and fitting a monogenic segregation model (3:1), was analyzed through BSA-seq, aiming at the identification of genomic regions/genes affecting male sterility. Four QTL regions were identified on chromosomes 4, 12, and 14. By analyzing the sequence around the highest pick on chromosome 14, a cytochrome P450 (CYP703A2) was identified, carrying a deleterious substitution (R/Q) fixed in the male sterile parent. A single dCAPS marker was developed around this SNP, allowing the discrimination between MS and MF genotypes within the population, suitable for applications in plant breeding programs. A 3D model of the protein was generated by homology modeling, revealing that the mutated amino acid is part of a highly conserved motif crucial for protein folding.

Bulked-segregant analysis by deep sequencing (BSA-seq) is a widely used method for mapping QTL (quantitative trait loci) due to its simplicity, speed, cost-effectiveness, and efficiency. However, the ability of BSA-seq to detect QTL is often limited by inappropriate experimental designs, as evidenced by numerous practical studies. Most BSA-seq studies have utilized small to medium-sized populations, with F2 populations being the most common choice. Nevertheless, theoretical studies have shown that using a large population with an appropriate pool size can significantly enhance the power and resolution of QTL detection in BSA-seq, with F3 populations offering notable advantages over F2 populations. To provide an experimental demonstration, we tested the power of BSA-seq to identify QTL controlling days from sowing to heading (DTH) in a 7200-plant rice F3 population in two environments, with a pool size of approximately 500. Each experiment identified 34 QTL, an order of magnitude greater than reported in most BSA-seq experiments, of which 23 were detected in both experiments, with 17 of these located near 41 previously reported QTL and eight cloned genes known to control DTH in rice. These results indicate that QTL mapping by BSA-seq in large F3 populations and multi-environment experiments can achieve high power, resolution, and reliability.

Fruit length (FL) is an important economical trait that affects fruit yield and appearance. Pumpkin (Cucurbita moschata Duch) contains a wealth genetic variation in fruit length. However, the natural variation underlying differences in pumpkin fruit length remains unclear. In this study, we constructed a F2 segregate population using KG1 producing long fruit and MBF producing short fruit as parents to identify the candidate gene for fruit length. By bulked segregant analysis (BSA-seq) and Kompetitive Allele-Specific PCR (KASP) approach of fine mapping, we obtained a 50.77 kb candidate region on chromosome 14 associated with the fruit length. Then, based on sequence variation, gene expression and promoter activity analyses, we identified a candidate gene (CmoFL1) encoding E3 ubiquitin ligase in this region may account for the variation of fruit length. One SNP variation in promoter of CmoFL1 changed the GT1CONSENSUS, and DUAL-LUC assay revealed that this variation significantly affected the promoter activity of CmoFL1. RNA-seq analysis indicated that CmoFL1 might associated with the cell division process and negatively regulate fruit length. Collectively, our work identifies an important allelic affecting fruit length, and provides a target gene manipulating fruit length in future pumpkin breeding. [ABSTRACT FROM AUTHOR]

The bacterial wilt disease caused by Ralstonia solanacearum is a significant threat to eggplant production. Breeding and promoting resistant varieties is one of the most effective methods to manage bacterial wilt. Conducting QTL (quantitative trait locus) mapping of resistant genes can substantially enhance the breeding of plant resistance to bacterial wilt. In this study, a population of 2200 F2 individuals derived from resistant and susceptible materials was utilized to establish extreme resistance and susceptibility pools. Following resequencing analysis of the parents and extreme pools, the QTL were examined using the DEEP-BSA software and QTLseqr R package (version 0.7.5.2). The results revealed that the detection of 10 QTL sites on chromosomes 5, 8, 9, and 11 by the five algorithms of the DEEP-BSA software. Additionally, the candidate region of 62 Mb–72 Mb on chromosome 5 was identified in all five algorithms of the DEEP-BSA software, as well as by the QTLseqr R package. Subsequent gene annotation uncovered 276 genes in the candidate region of 62 Mb–72 Mb on chromosome 5. Additionally, RNA-seq results indicated that only 13 genes had altered expression levels following inoculation with R. solanacearum in the resistant materials. Based on the expression levels, SMEL4_05g015980.1 and SMEL4_05g016110.1 were identified as candidate genes. Notably, SNP annotation identified a non-synonymous mutation in the exonic region of SMEL4_05g015980.1 and a variant in the promoter region of SMEL4_05g016110.1. The research findings have practical significance for the isolation of bacterial wilt resistance genes in eggplant and the development of resistance to bacterial wilt varieties in eggplant. [ABSTRACT FROM AUTHOR]

Fruit rind patterns are vital commercial quality traits in melon, in which the spotted or striped fruit rinds contribute to the commercial value of melon and can directly affect the choice of consumers. Although the spotted or non-spotted fruit rind pattern was studied in several cultivated melon accessions, the inheritance and regulating locus of this trait in wild melons are still unknown. Therefore, in this study, the inheritance and regulating loci of the green spotted fruit rind in a wild melon accession XNM020 were explored with F2 segregating populations derived from crossing between XNM020 and a cultivated melon XNM125. Segregating ratios of phenotypic data indicated that the green spotted fruit rind in XNM020 has a monogenic dominant inheritance. BSA-Seq showed that two potential genomic regions on chromosomes 4 (from 0.00 to 2.97 Mb) and 5 (from 0.00 to 2.34 Mb) regulate the formation of the green spotted fruit rind in wild melon XNM020. According to the annotations of polymorphic SNPs (single-nucleotide polymorphisms) and small InDels (insertions and deletions) in target genomic regions and the predicted gene functions, four genes MELO3C003316, MELO3C003375, MELO3C003388, and MELO3C014660 regulating chloroplast development or chlorophyll biosynthesis may be the best candidate genes. The results of this study enriched the inheritances of spotted fruit rinds in melon and also provided target genomic regions for marker-assisted selection breeding of melon focusing on fruit rinds. [ABSTRACT FROM AUTHOR]

Summary: Fruit firmness is an important trait in sweet cherry breeding because it directly positively influences fruit transportability, storage and shelf life. However, the underlying genes responsible and the molecular mechanisms that control fruit firmness remain unknown. In this study, we identified a candidate gene, PavSCPL, encoding a serine carboxypeptidase‐like protein with natural allelic variation, that controls fruit firmness in sweet cherry using map‐based cloning and functionally characterized PavSCPL during sweet cherry fruit softening. Genetic analysis revealed that fruit firmness in the 'Rainier' × 'Summit' F1 population was controlled by a single dominant gene. Bulked segregant analysis combined with fine mapping narrowed the candidate gene to a 473‐kb region (7418778–7 891 914 bp) on chromosome 6 which included 72 genes. The candidate gene PavSCPL, and a null allele harbouring a 5244‐bp insertion in the second exon that completely inactivated PavSCPL expression and resulted in the extra‐hard‐flesh phenotype, were identified by RNA‐sequencing analysis and gene cloning. Quantitative RT‐PCR analysis revealed that the PavSCPL expression level was increased with fruit softening. Virus‐induced gene silencing of PavSCPL enhanced fruit firmness and suppressed the activities of certain pectin‐degrading enzymes in the fruit. In addition, we developed functional molecular markers for PavSCPL and the Pavscpl5.2‐k allele that co‐segregated with the fruit firmness trait. Overall, this research identified a crucial functional gene for fruit firmness. The results provide insights into the genetic control and molecular mechanism of the fruit firmness trait and present useful molecular markers for molecular‐assisted breeding for fruit firmness in sweet cherry. [ABSTRACT FROM AUTHOR]

[Objective] To determine the position of tomato crack tolerance gene on chromosome. [Method] Tomato 14FP5 which is resistant to cracking and tomato 14FP26 which is easy to crack are used as test materials, cross the two to get F1 generation, F1 self-cross to get F2, DNA mixing pools with two extreme traits were constructed respectively, BSA-seq method was used to locate the location. [Result] BSA-seq was used for genome resequencing analysis of 2 extreme mixed pools of F2 population. The tomato anti-cracking gene was initially located on chromosome 5, located between 34 140 000 and 38 120 000 pb, the interval size is 3.98 Mb, a total of 38 genes were annotated in the candidate region, including 4 non-synonymous mutant genes. [Conclusion The results provide data support for the fine localization of tomato crack tolerance genes, and lay a foundation for tomato crack tolerance molecular breeding. [ABSTRACT FROM AUTHOR]

Copyright of Acta Agronomica Sinica is the property of Crop Science Society of China and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Bulked-segregant analysis by deep sequencing (BSA-seq) is a widely used method for mapping QTL (quantitative trait loci) due to its simplicity, speed, cost-effectiveness, and efficiency. However, the ability of BSA-seq to detect QTL is often limited by inappropriate experimental designs, as evidenced by numerous practical studies. Most BSA-seq studies have utilized small to medium-sized populations, with F2 populations being the most common choice. Nevertheless, theoretical studies have shown that using a large population with an appropriate pool size can significantly enhance the power and resolution of QTL detection in BSA-seq, with F3 populations offering notable advantages over F2 populations. To provide an experimental demonstration, we tested the power of BSA-seq to identify QTL controlling days from sowing to heading (DTH) in a 7200-plant rice F3 population in two environments, with a pool size of approximately 500. Each experiment identified 34 QTL, an order of magnitude greater than reported in most BSA-seq experiments, of which 23 were detected in both experiments, with 17 of these located near 41 previously reported QTL and eight cloned genes known to control DTH in rice. These results indicate that QTL mapping by BSA-seq in large F3 populations and multi-environment experiments can achieve high power, resolution, and reliability. [ABSTRACT FROM AUTHOR]

Lateral branching is an important agronomic trait in horticulture plants. The aim of this project was to reveal the genetic mechanism, map the gene localization, and predict candidate genes of watermelon lateral branching. An F2 segregating population was derived from a cross between the multibranched maternal inbred line M6 and the branchless paternal inbred line N7. Two DNA pools were constructed using 20 multibranched plants and 20 branchless plants from the F2 population. Whole-genome resequencing was performed for the DNA pools (25×) and the parents (30×) to identify the genomic region associated with lateral branching. Candidate genes were predicted based on the gene annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analyses; then, quantitative validation of these genes was performed. The results showed that the clean reads of four samples yielded 64,295,076 to 81,658,958 bp, with sufficient genome coverage and high quality. Based on single-nucleotide polymorphism and insertions/deletions association analyses, the candidate genes were mapped to a 2.01-Mb region on chromosome 4 (22,958,925–24,971,894 bp) containing 182 annotated genes. During the KEGG and GO enrichment analyses, these genes were annotated to 10 cellular components, 10 molecular functions, and 12 biological processes. Eight candidate genes responsible for the branchless phenotype in watermelon were identified: Cla97C04G076340, Cla97C04G075820, Cla97C04G076060, Cla97C04G076250, Cla97C04G076280, Cla97C04G076380, Cla97C04G076830, and Cla97C04G075950. These genes were involved in amino acid biosynthesis and catabolism, TCP transcription factor activity, and regulation of flower development. This study offers valuable insights into the molecular mechanisms governing the branchless phenotype in watermelon. These candidate genes serve as potential targets for gene cloning and marker-assisted selection of watermelon cultivars without lateral branches.

IntroductionFusarium head blight (FHB) has a large influence on both the yield and quality of wheat grain worldwide. Host resistance is the most effective method for controlling FHB, but unfortunately, very few genetic resources on FHB resistance are available; therefore, identifying novel resistance genes or quantitative trait loci (QTLs) is valuable. MethodsHere, a recombinant inbred line (RIL) population containing 451 lines derived from the cross L661/PI672538 was sown in four different environments (2019CZa, 2019CZb, 2021QL and 2021WJ).ResultsFive QTLs, consisting of two previously reported QTLs (FhbL693a and FhbL693b) and three new QTLs (FhbL693c, FhbL693d and FhbL693e), were identified. Further investigation revealed that FhbL693b, FhbL693c and FhbL693d could be detected in all four environments, and FhbL693a and FhbL693e were detected only in 2019CZb and 2021WJ, respectively. Among the QTLs, the greatest contribution (10.5%) to the phenotypic variation effect (PVE) was FhbL693d in 2021WJ, while the smallest (1.2%) was FhbL693e and FhbL693a in 2019CZb. The selection of 5Dindel-4 for FhbL693d, 4Aindel-7 for FhbL693c and 3Bindel-24 for FhbL693b decreased the number of damaged spikelets by 2.1, and a new line resistant to FHB named H140-2 was developed by marker-assisted selection (MAS). DiscussionThese results could help to further improve FHB resistance in the future.

Abstract Background Focusing on key indicators of drought resistance is highly important for quickly mining candidate genes related to drought resistance in cotton. Results In the present study, drought resistance was identified in drought resistance-related RIL populations during the flowering and boll stages, and multiple traits were evaluated; these traits included three key indicators: plant height (PH), single boll weight (SBW) and transpiration rate (Tr). Based on these three key indicators, three groups of extreme mixing pools were constructed for BSA-seq. Based on the mapping interval of each trait, a total of 6.27 Mb QTL intervals were selected on chromosomes A13 (3.2 Mb), A10 (2.45 Mb) and A07 (0.62 Mb) as the focus of this study. Based on the annotation information and qRT‒PCR analysis, three key genes that may be involved in the drought stress response of cotton were screened: GhF6'H1, Gh3AT1 and GhPER55. qRT‒PCR analysis of parental and extreme germplasm materials revealed that the expression of these genes changed significantly under drought stress. Cotton VIGS experiments verified the important impact of key genes on cotton drought resistance. Conclusions This study focused on the key indicators of drought resistance, laying the foundation for the rapid mining of drought-resistant candidate genes in cotton and providing genetic resources for directed molecular breeding of drought resistance in cotton.

Introduction: Branching angle is an essential trait in determining the planting density of rapeseed (Brassica napus L.) and hence the yield per unit area. However, the mechanism of branching angle formation in rapeseed is not well understood. Methods: In this study, two rapeseed germplasm with extreme branching angles were used to construct an F2 segregating population; then bulked segregant analysis sequencing (BSA-seq) and quantitative trait loci (QTL) mapping were utilized to localize branching anglerelated loci and combined with transcriptome sequencing (RNA-seq) and quantitative real-time PCR (qPCR) for candidate gene mining. Results and discussion: A branching angle-associated quantitative trait loci (QTL) was mapped on chromosome C3 (C3: 1.54-2.65 Mb) by combining BSAseq as well as traditional QTL mapping. A total of 54 genes had SNP/Indel variants within the QTL interval were identified. Further, RNA-seq of the two parents revealed that 12 of the 54 genes were differentially expressed between the two parents. Finally, we further validated the differentially expressed genes using qPCR and found that six of them presented consistent differential expression in all small branching angle samples and large branching angles, and thus were considered as candidate genes related to branching angles in rapeseed. Our results introduce new candidate genes for the regulation of branching angle formation in rapeseed, and provide an important reference for the subsequent exploration of its formation mechanism. [ABSTRACT FROM AUTHOR]

The soil-borne pathogen Plasmodiophora brassicae is the causal agent of clubroot, a major disease in Chinese cabbage (Brassica rapa ssp. pekinensis). The host's resistance genes often confer immunity to only specific pathotypes and may be rapidly overcome. Identification of novel clubroot resistance (CR) from germplasm sources is necessary. In this study, Bap246 was tested by being crossed with different highly susceptible B. rapa materials and showed recessive resistance to clubroot. An F2 population derived from Bap246 × Bac1344 was used to locate the resistance Quantitative Trait Loci (QTL) by Bulk Segregant Analysis Sequencing (BSA-Seq) and QTL mapping methods. Two QTL on chromosomes A01 (4.67–6.06 Mb) and A08 (10.42–11.43 Mb) were found and named Cr4Ba1.1 and Cr4Ba8.1, respectively. Fifteen and eleven SNP/InDel markers were used to narrow the target regions in the larger F2 population to 4.67–5.17 Mb (A01) and 10.70–10.84 Mb (A08), with 85 and 19 candidate genes, respectively. The phenotypic variation explained (PVE) of the two QTL were 30.97% and 8.65%, respectively. Combined with gene annotation, mutation site analysis, and real-time quantitative polymerase chain reaction (qRT-PCR) analysis, one candidate gene in A08 was identified, namely Bra020861. And an insertion and deletion (InDel) marker (co-segregated) named Crr1-196 was developed based on the gene sequence. Bra013275, Bra013299, Bra013336, Bra013339, Bra013341, and Bra013357 in A01 were the candidate genes that may confer clubroot resistance in Chinese cabbage. The resistance resource and the developed marker will be helpful in Brassica breeding programs. [ABSTRACT FROM AUTHOR]

Rice effective panicle is a major trait for grain yield and is affected by both the genetic tiller numbers and the early tillering vigor (ETV) traits to survive environmental adversities. The mechanism behind tiller bud formation has been well described, while the genes and the molecular mechanism underlying rice-regulating ETV traits are unclear. In this study, the candidate genes in regulating ETV traits have been sought by quantitative trait locus (QTL) mapping and bulk-segregation analysis by resequencing method (BSA-seq) conjoint analysis using rice backcross inbred line (BIL) populations, which were cultivated as late-season rice of double-cropping rice systems. By QTL mapping, seven QTLs were detected on chromosomes 1, 3, 4, and 9, with the logarithm of the odds (LOD) values ranging from 3.52 to 7.57 and explained 3.23% to 12.98% of the observed phenotypic variance. By BSA-seq analysis, seven QTLs on chromosomes 1, 2, 4, 5, 7, and 9 were identified using single-nucleotide polymorphism (SNP) and insertions/deletions (InDel) index algorithm and Euclidean distance (ED) algorithm. The overlapping QTL resulting from QTL mapping and BSA-seq analysis was shown in a 1.39 Mb interval on chromosome 4. In the overlap interval, six genes, including the functional unknown genes Os04g0455650, Os04g0470901, Os04g0500600, and ethylene-insensitive 3 (Os04g0456900), sialyltransferase family domain containing protein (Os04g0506800), and ATOZI1 (Os04g0497300), showed the differential expression between ETV rice lines and late tillering vigor (LTV) rice lines and have a missense base mutation in the genomic DNA sequences of the parents. We speculate that the six genes are the candidate genes regulating the ETV trait in rice, which provides a research basis for revealing the molecular mechanism behind the ETV traits in rice. [ABSTRACT FROM AUTHOR]

The yield of soybean is positively correlated with the duration of reproductive growth, which delays the aging of leaves after flowering, enhances their physiological performance, and supports growing plants with heavier grains. Leaf yellowing is one of the distinctive features of plant aging. Studies on leaf yellowing at the late stage of soybean drum grain have rarely been reported. The early yellowing feature of soybean late tympanic leaves was controlled by a single recessive nuclear gene, according to the genetic analysis of the hybrid of the early yellowing mutant ly and wild-type ofc in this study. Using Molecular Marker to Map-Based Cloning, a 2.23 Mb preliminary localization interval on chromosome 19 was obtained. The interval was shortened to 1.75 Mb and contained 219 genes. When this interval was combined with RNA-Seq analysis, 12 candidate genes were detected, including 4 SNP variant genes and 8 differentially expressed genes. The findings of this study provides the framework for the cloning of genes that cause aging and yellowing during the filling later period in soybean. [ABSTRACT FROM AUTHOR]

Plant photosynthesis is mainly dependent on leaf color, and this has an impact on yield. Mutants lacking in chlorophyll have been analyzed to gain insight into the genetic processes involved in photosynthesis, chloroplast development, and chlorophyll metabolism. A yellow-to-lethal mutant, ytl, was selected from the M6 generation of the 60Coγ ray irradiation-treated soybean cultivar Nannong 1138-2. The mutant exhibited reduced chlorophyll content, with the thylakoid structure disrupted. Segregation of the cross between Williams 82 (W82) and ytl indicated that a recessive allele controlled yellow-to-lethal traits. The bulked-segregant analysis (BSA)-Seq method performed preliminary mapping, followed by simple sequence repeat (SSR) marker validation and further mapping. The candidate gene was mapped to a 418 Kb region containing 53 genes. High-throughput sequencing and first-generation sequencing results showed a two bp deletion in the second exon of Glyma.08g106500, leading to a frameshift mutation in ytl. As a promising candidate gene, Glyma.08g106500 encoded a chloroplast-localized pentatricopeptide repeat (PPR) domain-containing protein involved in the assembly of chloroplast proteins. These results will contribute to cloning the mutant ytl gene and provide insight into the regulatory processes controlling photosynthesis and chloroplast development and growth in soybean. [ABSTRACT FROM AUTHOR]

Soybean is a primary source of plant-based oil and protein for human diets. Seed size and weight are important agronomic traits that significantly influence soybean yield. Despite their importance, the genetic mechanisms underlying soybean seed size and weight remain to be fully elucidated. In order to identify additional, major quantitative trait loci (QTL) associated with seed size and weight, we developed segregating populations by crossing a large-seeded soybean variety “Kebaliang” with a small-seeded soybean variety “SUZUMARU”. We evaluated seed length, width, thickness, and hundred-seed weight across two generations, F4 and F5, in 2022 and 2023. Employing bulked segregate analysis with whole-genome resequencing (BSA-seq), we detected 18 QTLs in the F4 population and 12 QTLs in the F5 population. Notably, six QTLs showed high stability between the two generations, with five derived from two pleiotropic loci (qSS4-1 and qSS20-1) and one specific to seed width (qSW14-1). Further validation and refinement of these loci were carried out through linkage mapping using molecular markers in the F5 population. Additionally, we identified 18 candidate genes within these stable loci and analyzed their sequence variations and expression profiles. Together, our findings offered a foundational reference for further soybean seed size research and unveiled novel genetic loci and candidate genes that could be harnessed for the genetic enhancement of soybean production.

Plant height, as a crucial component of plant architecture, exerts a significant influence on rapeseed (Brassica napus L.) lodging resistance, photosynthetic efficiency, yield, and mechanized harvest level. A previous study identified dwarf rapeseed LSW2018. In this study, LSW2018 (dwarf parent (PD)) was crossed with 389 (high parent (PH)) to establish the F2 population, and 30 extremely dwarf (bulk–D) and high (bulk–H) plants in the F2 population were respectively selected to construct two bulked DNA pools. Whole–genome sequencing and variation analysis (BSA–seq) were performed on these four DNA pools (PD, PH, bulk–D, and bulk–H). The BSA–seq results revealed that the genomic region responsible for the dwarf trait spanned from 19.30 to 22.19 Mb on chromosome A03, with a length of 2.89 Mb. After fine mapping with simple sequence repeat (SSR) markers, the gene was narrowed to a 0.71 Mb interval. Within this region, a total of 113 genes were identified, 42 of which contained large–effect variants. According to reference genome annotation and qRT–PCR analysis, there are 17 differentially expressed genes in this region between high and dwarf individuals. This study preliminarily reveals the genetic basis of LSW2018 dwarfing and provides a theoretical foundation for the molecular marker–assisted breeding of dwarf rapeseed.

Fruit size is a crucial agronomic trait in bottle gourd, impacting both yield and utility. Despite its significance, the regulatory mechanism governing fruit size in bottle gourd remains largely unknown. In this study, we used bottle gourd (small-fruited H28 and large-fruited H17) parent plants to measure the width and length of fruits at various developmental stages, revealing a single ‘S’ growth curve for fruit expansion. Paraffin section observations indicated that both cell number and size significantly influence bottle gourd fruit size. Through bulked segregant analysis and combined genotype–phenotype analysis, the candidate interval regulating fruit size was pinpointed to 17,747,353 bp–18,185,825 bp on chromosome 9, encompassing 0.44 Mb and including 44 genes. Parental fruits in the rapid expansion stage were subjected to RNA-seq, highlighting that differentially expressed genes were mainly enriched in pathways related to cell wall biosynthesis, sugar metabolism, and hormone signaling. Transcriptome and resequencing analysis, combined with gene function annotation, identified six genes within the localized region as potential regulators of fruit size. This study not only maps the candidate interval of genes influencing fruit size in bottle gourd through forward genetics, but also offers new insights into the potential molecular mechanisms underlying this trait through transcriptome analysis.

Since the official release of the stand-alone bioinformatics toolkit TBtools in 2020, its superior functionality in data analysis has been demonstrated by its widespread adoption by many thousands of users and references in more than 5000 academic articles. Now, TBtools is a commonly used tool in biological laboratories. Over the past 3 years, thanks to invaluable feedback and suggestions from numerous users, we have optimized and expanded the functionality of the toolkit, leading to the development of an upgraded version—TBtools-II. In this upgrade, we have incorporated over 100 new features, such as those for comparative genomics analysis, phylogenetic analysis, and data visualization. Meanwhile, to better meet the increasing needs of personalized data analysis, we have launched the plugin mode, which enables users to develop their own plugins and manage their selection, installation, and removal according to individual needs. To date, the plugin store has amassed over 50 plugins, with more than half of them being independently developed and contributed by TBtools users. These plugins offer a range of data analysis options including co-expression network analysis, single-cell data analysis, and bulked segregant analysis sequencing data analysis. Overall, TBtools is now transforming from a stand-alone software to a comprehensive bioinformatics platform of a vibrant and cooperative community in which users are also developers and contributors. By promoting the theme "one for all, all for one", we believe that TBtools-II will greatly benefit more biological researchers in this big-data era. An upgraded version of TBtools (TBtools-II) is introduced along with over 100 new functions and a novel plugin mode featuring tens of useful plugins. In TBtools-II, users can not only explore advanced functionalities but can also contribute to the development of plugins that cater to more specific data analysis purposes. By embracing the "one for all, all for one" theme, TBtools-II aims to evolve into a versatile bioinformatics platform capable of accommodating a wide range of personalized data analysis requirements. [ABSTRACT FROM AUTHOR]