Transcriptomics and virus-induced gene silencing identify defence-related genes during Ralstonia solanacearum infection in resistant and susceptible tobacco.

Bacterial wilt (BW) caused by Ralstonia solanacearum is a globally prevalent bacterial soil-borne disease. In this study, transcriptome sequencing were subjected to roots after infection with the R. solanacearum in the resistant and susceptible tobacco variety. DEGs that responded to R. solanacearum infection in both resistant and susceptible tobacco contributed to pectinase and peroxidase development and were enriched in plant hormone signal transduction, signal transduction and MAPK signalling pathway KEGG terms. Core DEGs in the resistant tobacco response to R. solanacearum infection were enriched in cell wall, membrane, abscisic acid and ethylene terms. qRT-PCR indicated that Nitab4.5_0004899g0110, Nitab4.5_0004234g0080 and Nitab4.5_0001439g0050 contributed to the response to R. solanacearum infection in different resistant and susceptible tobacco. Silencing the p450 gene Nitab4.5_0001439g0050 reduced tobacco resistance to bacterial wilt. These results improve our understanding of the molecular mechanism of BW resistance in tobacco and solanaceous plants.

PCMDB: a curated and comprehensive resource of plant cell markers.

The advent of single-cell sequencing opened a new era in transcriptomic and genomic research. To understand cell composition using single-cell studies, a variety of cell markers have been widely used to label individual cell types. However, the specific database of cell markers for use by the plant research community remains very limited. To overcome this problem, we developed the Plant Cell Marker DataBase (PCMDB, http://www.tobaccodb.org/pcmdb/), which is based on a uniform annotation pipeline. By manually curating over 130 000 research publications, we collected a total of 81 117 cell marker genes of 263 cell types in 22 tissues across six plant species. Tissue- and cell-specific expression patterns can be visualized using multiple tools: eFP Browser, Bar, and UMAP/TSNE graph. The PCMDB also supports several analysis tools, including SCSA and SingleR, which allows for user annotation of cell types. To provide information about plant species currently unsupported in PCMDB, potential marker genes for other plant species can be searched based on homology with the supported species. PCMDB is a user-friendly hierarchical platform that contains five built-in search engines. We believe PCMDB will constitute a useful resource for researchers working on cell type annotation and the prediction of the biological function of individual cells.

A dimensionality-reduction genomic prediction method without direct inverse of the genomic relationship matrix for large genomic data.

KEY MESSAGE: A new genomic prediction method (RHPP) was developed via combining randomized Haseman-Elston regression (RHE-reg), PCR based on genomic information of core population, and preconditioned conjugate gradient (PCG) algorithm. Computational efficiency is becoming a hot issue in the practical application of genomic prediction due to the large number of data generated by the high-throughput genotyping technology. In this study, we developed a fast genomic prediction method RHPP via combining randomized Haseman-Elston regression (RHE-reg), PCR based on genomic information of core population, and preconditioned conjugate gradient (PCG) algorithm. The simulation results demonstrated similar prediction accuracy between RHPP and GBLUP, and significantly higher computational efficiency of the former with the increase of individuals. The results of real datasets of both bread wheat and loblolly pine demonstrated that RHPP had a similar or better predictive accuracy in most cases compared with GBLUP. In the future, RHPP may be an attractive choice for analyzing large-scale and high-dimensional data.

PLncDB V2.0: a comprehensive encyclopedia of plant long noncoding RNAs.

Long noncoding RNAs (lncRNAs) are transcripts longer than 200 nucleotides with little or no protein coding potential. The expanding list of lncRNAs and accumulating evidence of their functions in plants have necessitated the creation of a comprehensive database for lncRNA research. However, currently available plant lncRNA databases have some deficiencies, including the lack of lncRNA data from some model plants, uneven annotation standards, a lack of visualization for expression patterns, and the absence of epigenetic information. To overcome these problems, we upgraded our Plant Long noncoding RNA Database (PLncDB, http://plncdb.tobaccodb.org/), which was based on a uniform annotation pipeline. PLncDB V2.0 currently contains 1 246 372 lncRNAs for 80 plant species based on 13 834 RNA-Seq datasets, integrating lncRNA information from four other resources including EVLncRNAs, RNAcentral and etc. Expression patterns and epigenetic signals can be visualized using multiple tools (JBrowse, eFP Browser and EPexplorer). Targets and regulatory networks for lncRNAs are also provided for function exploration. In addition, PLncDB V2.0 is hierarchical and user-friendly and has five built-in search engines. We believe PLncDB V2.0 is useful for the plant lncRNA community and data mining studies and provides a comprehensive resource for data-driven lncRNA research in plants.

Nitrogen application and differences in leaf number retained after topping affect the tobacco (Nicotiana tabacum) transcriptome and metabolome.

BACKGROUND: Agronomic treatments such as the application of nitrogen fertilizer and topping (removal of the inflorescence and top leaves) cause substantial changes in plant metabolism. To explore these changes, we conducted comparative transcriptomic and metabolomic analyses of leaves collected from four positions along the stem on plants exposed to two nitrogen doses and with different numbers of leaves retained after topping in tobacco (Nicotiana tabacum). RESULTS: We identified 13,330 unique differentially expressed genes and 32 differentially abundant metabolites. Through RNA-seq and WGCNA analyze, we constructed 2 co-expression networks (green and blue) highly correlation to N application and leaf number retained, predicted a hub gene NtGER3 may play an important role in N metabolism related to amino acid (cysteine) through CK pathway in tobacco leaves, NtARFs may participated in modulating the auxin signal and N in bottom leaves and NtRAP2.12 as key gene involved in N regulation by ethylene pathway. What's more, our data prove C/N transformation and balance affect the "source - flow - sink" redistribution and remobilization in tobacco during growth and development process. CONCLUSIONS: Overall, this comparative transcriptomics study provides novel insight into the complex molecular mechanisms underlying plant responses to different levels of nitrogen application and the number of leaves remaining after topping in plants.

Genome-wide association study revealed TaHXK3-2A as a candidate gene controlling stomatal index in wheat seedlings.

Stomata are important channels for the control of gas exchange between plants and the atmosphere. To examine the genetic architecture of wheat stomatal index, we performed a genome-wide association study (GWAS) using a panel of 539 wheat accessions and 450 678 polymorphic single nucleotide polymorphisms (SNPs) that were detected using wheat-specific 660K SNP array. A total of 130 SNPs were detected to be significantly associated with stomatal index in both leaf surfaces of wheat seedlings. These significant SNPs were distributed across 16 chromosomes and involved 2625 candidate genes which participate in stress response, metabolism and cell/organ development. Subsequent bulk segregant analysis (BSA), combined with GWAS identified one major haplotype on chromosome 2A, that is responsible for stomatal index on the abaxial leaf surface. Candidate gene association analysis revealed that genetic variation in the promoter region of the hexokinase gene TaHXK3-2A was significantly associated with the stomatal index. Moreover, transgenic analysis confirmed that TaHXK3-2A overexpression in wheat decreased the size of leaf pavement cells but increased stomatal density through the glucose metabolic pathway, resulting in drought sensitivity among TaHXK3-2A transgenic lines due to an increased transpiration rate. Taken together, these results provide valuable insights into the genetic control of the stomatal index in wheat seedlings.

Comparative transcriptomic analysis of seed coats with high and low lignin contents reveals lignin and flavonoid biosynthesis in Brassica napus.

BACKGROUND: Brassica napus L. (2n = 38, AACC) is one of the most important oil crops and sources of protein for animal feed worldwide. Lignin is a large molecule aromatic polymer and a major cell wall component. However, lignin in the seed coat reduces the availability and restricts the development of rapeseed cake. Therefore, it is critical to reduce the lignin content of the seed coat. Here, high-lignin (H-lignin) and low-lignin (L-lignin) content recombinant inbred lines (RILs) were selected from an RIL population for analysis. RESULTS: The cross-section results indicated that the seed coat of the H-lignin lines was thicker than that of the L-lignin lines, especially the palisade layer. The seed coats and embryos at 35, 40 and 46 days after flowering (DAF) were subjected to RNA sequencing (RNA-Seq), and the expression of the BnPAL and BnC4H gene families in the lignin pathway was significantly higher in the H-lignin seed coat than in the L-lignin seed coat. The Bn4CL gene family also showed this trend. In addition, among the genes related to plant hormone synthesis, BnaC02g01710D was upregulated and BnaA07g11700D and BnaC09g00190D were downregulated in H-lignin lines. Some transcription factors were upregulated, such as BnNAC080, BnNAC083, BnMYB9, BnMYB9-1, BnMYB60 and BnMYB60-1, while BnMYB91 was downregulated in H-lignin lines. Moreover, most genes of the flavonoid pathway, such as BnCHS and BnDFR, were strongly expressed in H-lignin seed coat. CONCLUSIONS: In Our study, some key genes such as hormone synthesis genes, transcription factors and miRNAs related to lignin and flavonoid biosynthesis were identified. A regulatory model of B. napus seed coat lignin was proposed. These results provide new insight into lignin and flavonoid biosynthesis in B. napus.

A large-scale genomic association analysis identifies the candidate causal genes conferring stripe rust resistance under multiple field environments.

The incorporation of resistance genes into wheat commercial varieties is the ideal strategy to combat stripe or yellow rust (YR). In a search for novel resistance genes, we performed a large-scale genomic association analysis with high-density 660K single nucleotide polymorphism (SNP) arrays to determine the genetic components of YR resistance in 411 spring wheat lines. Following quality control, 371 972 SNPs were screened, covering over 50% of the high-confidence annotated gene space. Nineteen stable genomic regions harbouring 292 significant SNPs were associated with adult-plant YR resistance across nine environments. Of these, 14 SNPs were localized in the proximity of known loci widely used in breeding. Obvious candidate SNP variants were identified in certain confidence intervals, such as the cloned gene Yr18 and the major locus on chromosome 2BL, despite a large extent of linkage disequilibrium. The number of causal SNP variants was refined using an independent validation panel and consideration of the estimated functional importance of each nucleotide polymorphism. Interestingly, four natural polymorphisms causing amino acid changes in the gene TraesCS2B01G513100 that encodes a serine/threonine protein kinase (STPK) were significantly involved in YR responses. Gene expression and mutation analysis confirmed that STPK played an important role in YR resistance. PCR markers were developed to identify the favourable TraesCS2B01G513100 haplotype for marker-assisted breeding. These results demonstrate that high-resolution SNP-based GWAS enables the rapid identification of putative resistance genes and can be used to improve the efficiency of marker-assisted selection in wheat disease resistance breeding.

Systematic study of the stress-responsive Rboh gene family in Nicotiana tabacum: Genome-wide identification, evolution and role in disease resistance.

Plant respiratory burst oxidase homolog (Rboh) gene family encodes the key enzymatic subunits of reactive oxygen species (ROS) production pathways, and play crucial role in plant signaling, development and stress responses. In present work, twenty genes were identified in Nicotiana tabacum Rboh family (NtabRboh) and classified into four phylogenetic groups (I-IV). Fourteen NtabRboh genes were positioned on ten chromosomes (i.e., Ch1, 2, 4, 7-11, 14 and 21), and six scaffolds. Synteny and evolutionary analysis showed that most of the NtabRboh genes have evolved from the genomes of the ancestor species (N. tomentosiformis and N. sylvestris), which afterwards expanded through duplication events. The promoter regions of the NtabRboh genes contained numerous cis-acting regulatory elements for hormones, plant growth, and different biotic and abiotic factors. The NtabRbohF gene transcript comprised target sites for wounding and stress responsive microRNAs: nta-miR166a-d, g and h. The transcript abundance of NtabRboh genes in different tissues reflected their important for plant growth and organ development in tobacco. RT-qPCR-assays demonstrated that the expression of NtabRboh genes are regulated by viral and bacterial pathogens, drought, cold and cadmium stress. The expression levels NtabRbohA, B and C were significantly up-regulated in "black shank and tobacco mosaic virus-inoculated susceptible and transgenic tobacco cultivars, showing that these genes play important roles in disease resistance.

Comparative transcriptome and metabolomic profiling reveal the complex mechanisms underlying the developmental dynamics of tobacco leaves.

Although the leaf is the most important photosynthetic organ in most plants, many of the molecular mechanisms underlying leaf developmental dynamics remain to be explored. To better understand the transcriptional regulatory mechanisms involved in leaf development, we conducted comparative transcriptomic and metabolomic analysis of leaves from seven positions on tobacco (Nicotiana tabacum) plants. A total of 35,622 unique differentially expressed genes and 79 metabolites were identified. A time-series expression analysis detected two interesting transcriptional profiles, one comprising 10,197 genes that displayed continual up-regulation during leaf development and another comprising 4696 genes that displayed continual down-regulation. Combining these data with co-expression network results identified four important regulatory networks involved in photorespiration and the tricarboxylic acid cycle; these networks may regulate carbon/nitrogen balance during leaf development. We also found that the transcription factor NtGATA5 acts as a hub associated with C and N metabolism and chloroplast development during leaf development through regulation of phytohormones. Furthermore, we investigated the transcriptional dynamics of genes involved in the auxin, cytokinin, and jasmonic acid biosynthesis and signaling pathways during tobacco leaf development. Overall, our study greatly expands the understanding of the regulatory network controlling developmental dynamics in plant leaves.

Association Analysis Identifies New Loci for Resistance to Chinese Yr26-Virulent Races of the Stripe Rust Pathogen in a Diverse Panel of Wheat Germplasm.

Stripe rust caused by Puccinia striiformis f. sp. tritici (Pst) is one of the most destructive fungal diseases of wheat worldwide. The expanding Yr26-virulent Pst race (V26) group overcomes almost all currently deployed resistance genes in China and has continued to accumulate new virulence. Investigating the genetic architecture of stripe rust resistance in common wheat is an important basis for a successful utilization of resistance in breeding programs. A panel of 410 exotic wheat germplasms was used for characterizing new stripe rust resistance loci. This panel was genotyped using high-density wheat 660K single-nucleotide polymorphism (SNP) array, and phenotypic evaluation of seedlings for stripe rust resistance was performed using multiple Pst races. Thirty-five loci conferring resistance were identified through genome-wide association mapping, and explained phenotypic variances ranged from 53 to 75%. Of these, 14 were colocated in the proximity of the known loci, including cataloged Yr genes Yr9, Yr10, Yr26, Yr33, Yr47, Yr56, Yr57, Yr64, Yr67, Yr72, and Yr81 and three temporarily designated as YrCen, YrNP63, and YrRC detected in our quantitative trait locus (QTL) mapping studies. Seven of them (Yr9, Yr10, Yr24/26, Yr81, YrCEN, YrNP63, and YrRC) were confirmed by molecular detection or genetic analysis. New loci that were identified to be different from reported Yr genes need further confirmation. Nine QTL with significantly large phenotypic effect on resistance to all tested races were considered as major loci for effective resistance. The identified loci enrich our stripe rust resistance gene pool, and the linked SNPs should be useful for marker-assisted selection in breeding programs.

Genome-Scale Analysis of Homologous Genes among Subgenomes of Bread Wheat (Triticum aestivum L.).

Determining the distribution and correspondence of genome-scale homologous genes in wheat are effective ways to uncover chromosome rearrangement that has occurred during crop evolution and domestication, which can contribute to improvements in crop breeding. High-resolution and comprehensive analysis of the wheat genome by the International Wheat Genome Sequencing Consortium (IWGSC) revealed a total of 88,733 high-confidence homologous genes of four major types (1:1:1, 1:1:0, 0:1:1 and 1:0:1) among the A, B and D subgenomes of wheat. This data was used to compare homologous gene densities among chromosomes, clarify their distribution and correspondence relationship, and compare their functional enrichment. The average density of 1:1:1 homologous genes was about 10 times more than the density of the other three types of homologous genes, although the homologous gene densities of the various chromosomes were similar within each homologous type. Three regions of exceptional density were detected in 1:1:1 homologous genes, the isolate peak on the tail of chromosome 4A, and the desert regions at the start of chromosome 7A and 7D. The correspondence between homologous genes of the wheat subgenomes demonstrated translocation between the tail segments of chromosome 4A and 5A, and the inversion of the segment of original 5A and 7B into the tail of 4A. The homologous genes on the inserting segments of 5A and 7B to 4A were highly enriched in nitrogen, primary metabolite and small molecular metabolism processes, compared with genes on other regions of the original 4A chromosome. This study provides a refined genome-scale reference of homologous genes for wheat molecular research and breeding, which will help to broaden the application of the wheat genome and can be used as a template for research on other polyploid plants.

Genetic architecture of wheat stripe rust resistance revealed by combining QTL mapping using SNP-based genetic maps and bulked segregant analysis.

KEY MESSAGE: A major stripe rust resistance QTL was mapped to a 0.4 centimorgan (cM) genetic region on the long arm of chromosome 7B, using combined genome-wide linkage mapping and bulk segregant analysis. The German winter wheat cv. Centrum has displayed high levels of adult plant stripe rust resistance (APR) in field environments for many years. Here, we used the combined genome-wide linkage mapping and pool-extreme genotyping to characterize the APR resistance. One hundred and fifty-one F2:7 recombinant inbred lines derived from a cross between susceptible landrace Mingxian 169 and Centrum were evaluated for stripe rust resistance in multiple environments and genotyped by the wheat 35K single nucleotide polymorphism (SNP) array. Three stable quantitative trait loci (QTL) were identified using QTL analysis across five field environments. To saturate the major QTL, the wheat 660K SNP array was also used to genotype bulked extremes. A major QTL named QYrcen.nwafu-7BL from Centrum was mapped in a 0.4 cM genetic interval flanking by AX-94556751 and AX-110366788 across a 2 Mb physical genomic region, explaining 19.39-42.81% of the total phenotypic variation. It is likely a previously uncharacterized QTL based on pedigree analysis, reaction response, genotyping data and map comparison. The SNP markers closely linked with QYrcen.nwafu-7BL were converted to KASP markers and validated in a subset of 120 wheat lines. A 211 F2 breeding population from a cross of an elite cultivar Xinong 979 with Centrum were developed for marker-based selection. Three selected lines with desirable agronomic traits and the positive alleles of both KASP markers showed acceptable resistance which should be used as resistance donors in wheat breeding programs. The other QTL QYrcen.nwafu-1AL and QYrcen.nwafu-4AL with additive effects could enhance the level of resistance conferred by QYrcen.nwafu-7BL.

A major QTL co-localized on chromosome 6BL and its epistatic interaction for enhanced wheat stripe rust resistance.

KEY MESSAGE: Co-localization of a major QTL for wheat stripe rust resistance to a 3.9-cM interval on chromosome 6BL across both populations and another QTL on chromosome 2B with epistatic interaction. Cultivars with diverse resistance are the optimal strategy to minimize yield losses caused by wheat stripe rust (Puccinia striiformis f. sp. tritici). Two wheat populations involving resistant wheat lines P10078 and Snb"S" from CIMMYT were evaluated for stripe rust response in multiple environments. Pool analysis by Wheat660K SNP array showed that the overlapping interval on chromosome 6B likely harbored a major QTL between two populations. Then, linkage maps were constructed using KASP markers, and a co-localized locus with large effect on chromosome 6BL was detected using QTL analysis in both populations. The coincident QTL, named QYr.nwafu-6BL.2, explained 59.7% of the phenotypic maximum variation in the Mingxian 169 × P10078 and 52.5% in the Zhengmai 9023 × Snb"S" populations, respectively. This co-localization interval spanning 3.9 cM corresponds to ~ 30.5-Mb genomic region of the newest common wheat reference genome (IWGSC RefSeq v.1.0). In addition, another QTL was also detected on chromosome 2B in Zhengmai 9023 × Snb"S" population and it can accelerate expression of QYr.nwafu-6BL.2 to enhance resistance with epistatic interaction. Allowing for Pst response, marker genotypes, pedigree analysis and relative genetic distance, QYr.nwafu-6BL.2 is likely to be a distinct adult plant resistance QTL. Haplotype analysis of QYr.nwafu-6BL.2 revealed specific SNPs or alleles in the target region from a diversity panel of 176 unrelated wheat accessions. This QTL region provides opportunity for further map-based cloning, and haplotypes analysis enables pyramiding favorable alleles into commercial cultivars by marker-assisted selection.

Genome-Wide Linkage Mapping Reveals Stripe Rust Resistance in Common Wheat (Triticum aestivum) Xinong1376.

Stripe rust, also known as yellow rust, is a significant threat to wheat yield worldwide. Adult plant resistance (APR) is the preferred way to obtain durable protection. Chinese winter wheat cultivar Xinong1376 has maintained acceptable APR to stripe rust in field environments. To characterize APR in this cultivar, 190 F10 recombinant inbred lines (RILs) developed from Xiaoyan81 × Xinong1376 were evaluated for infection type and disease severity in fields either artificially or naturally inoculated. The population along with parents were genotyped using the Illumina 90K single-nucleotide polymorphism arrays. Six quantitative trait loci (QTL) were detected using the inclusive composite interval mapping method. QYr.nwafu-4AL and QYr.nwafu-6BL.3 conferred stable resistance in all environments, and likely corresponded to a gene-rich region on the long arm of chromosomes 4A and 6B. QYr.nwafu-5AL, QYr.nwafu-5BL, QYr.nwafu-3BL.1, and QYr.nwafu-3BL.2 were detected only in some environments but enhanced the level of resistance conferred by QYr.nwafu-4AL and QYr.nwafu-6BL.3. Kompetitive allele-specific PCR (KASP) markers developed for QYr.nwafu-4AL and QYr.nwafu-6BL.3 were confirmed in a subset of RILs and 133 wheat genotypes. The QTL on 4AL and 6BL with their linked KASP markers would be useful for marker-assisted selection to improve stripe rust resistance in breeding programs.

Rapid identification of an adult plant stripe rust resistance gene in hexaploid wheat by high-throughput SNP array genotyping of pooled extremes.

KEY MESSAGE: High-throughput SNP array analysis of pooled extreme phenotypes in a segregating population by KASP marker genotyping permitted rapid, cost-effective location of a stripe rust resistance QTL in wheat. German wheat cultivar "Friedrichswerther" has exhibited high levels of adult plant resistance (APR) to stripe rust in field environments for many years. F2:3 lines and F6 recombinant inbred line (RILs) populations derived from a cross between Friedrichswerther and susceptible landrace Mingxian 169 were evaluated in the field in 2013, 2016 and 2017. Illumina 90K iSelect SNP arrays were used to genotype bulked extreme pools and parents; 286 of 1135 polymorphic SNPs were identified on chromosome 6B. Kompetitive Allele-Specific PCR (KASP) markers were used to verify the chromosome region associated with the resistance locus. A linkage map was constructed with 18 KASP-SNP markers, and a major effect QTL was identified within a 1.4 cM interval flanked by KASP markers IWB71602 and IWB55937 in the region 6BL3-0-0.36. The QTL, named QYr.nwafu-6BL, was stable across environments, and explained average 54.4 and 47.8% of the total phenotypic variation in F2:3 lines and F6 RILs, respectively. On the basis of marker genotypes, pedigree analysis and relative genetic distance QYr.nwafu-6BL is likely to be a new APR QTL. Combined high-throughput SNP array genotyping of pooled extremes and validation by KASP assays lowers sequencing costs compared to genome-wide association studies with SNP arrays, and more importantly, permits rapid isolation of major effect QTL in hexaploid wheat as well as improving accuracy of mapping in the QTL region. QYr.nwafu-6BL with flanking KASP markers developed and verified in a subset of 236 diverse lines can be used in marker-assisted selection to improve stripe rust resistance in breeding programs.

Comparative genome-wide mapping versus extreme pool-genotyping and development of diagnostic SNP markers linked to QTL for adult plant resistance to stripe rust in common wheat.

KEY MESSAGE: A major stripe rust resistance QTL on chromosome 4BL was localized to a 4.5-Mb interval using comparative QTL mapping methods and validated in 276 wheat genotypes by haplotype analysis. CYMMIT-derived wheat line P10103 was previously identified to have adult plant resistance (APR) to stripe rust in the greenhouse and field. The conventional approach for QTL mapping in common wheat is laborious. Here, we performed QTL detection of APR using a combination of genome-wide scanning and extreme pool-genotyping. SNP-based genetic maps were constructed using the Wheat55 K SNP array to genotype a recombinant inbred line (RIL) population derived from the cross Mingxian 169 × P10103. Five stable QTL were detected across multiple environments. A fter comparing SNP profiles from contrasting, extreme DNA pools of RILs six putative QTL were located to approximate chromosome positions. A major QTL on chromosome 4B was identified in F2:4 contrasting pools from cross Zhengmai 9023 × P10103. A consensus QTL (LOD = 26-40, PVE = 42-55%), named QYr.nwafu-4BL, was defined and localized to a 4.5-Mb interval flanked by SNP markers AX-110963704 and AX-110519862 in chromosome arm 4BL. Based on stripe rust response, marker genotypes, pedigree analysis and mapping data, QYr.nwafu-4BL is likely to be a new APR QTL. The applicability of the SNP-based markers flanking QYr.nwafu-4BL was validated on a diversity panel of 276 wheat lines. The additional minor QTL on chromosomes 4A, 5A, 5B and 6A enhanced the level of resistance conferred by QYr.nwafu-4BL. Marker-assisted pyramiding of QYr.nwafu-4BL and other favorable minor QTL in new wheat cultivars should improve the level of APR to stripe rust.

SNP-based pool genotyping and haplotype analysis accelerate fine-mapping of the wheat genomic region containing stripe rust resistance gene Yr26.

KEY MESSAGE: NGS-assisted super pooling emerging as powerful tool to accelerate gene mapping and haplotype association analysis within target region uncovering specific linkage SNPs or alleles for marker-assisted gene pyramiding. Conventional gene mapping methods to identify genes associated with important agronomic traits require significant amounts of financial support and time. Here, a single nucleotide polymorphism (SNP)-based mapping approach, RNA-Seq and SNP array assisted super pooling analysis, was used for rapid mining of a candidate genomic region for stripe rust resistance gene Yr26 that has been widely used in wheat breeding programs in China. Large DNA and RNA super-pools were genotyped by Wheat SNP Array and sequenced by Illumina HiSeq, respectively. Hundreds of thousands of SNPs were identified and then filtered by multiple filtering criteria. Among selected SNPs, over 900 were found within an overlapping interval of less than 30 Mb as the Yr26 candidate genomic region in the centromeric region of chromosome arm 1BL. The 235 chromosome-specific SNPs were converted into KASP assays to validate the Yr26 interval in different genetic populations. Using a high-resolution mapping population (> 30,000 gametes), we confined Yr26 to a 0.003-cM interval. The Yr26 target region was anchored to the common wheat IWGSC RefSeq v1.0 and wild emmer WEWSeq v.1.0 sequences, from which 488 and 454 kb fragments were obtained. Several candidate genes were identified in the target genomic region, but there was no typical resistance gene in either genome region. Haplotype analysis identified specific SNPs linked to Yr26 and developed robust and breeder-friendly KASP markers. This integration strategy can be applied to accelerate generating many markers closely linked to target genes/QTL for a trait of interest in wheat and other polyploid species.

[Identification and expression analysis of TCP family members in tobacco (Nicotiana tabacum L.)].

TCP family as plant specific transcription factor, plays an important role in different aspects of plant development. In order to screen TCP family members in tobacco, the homologous sequences of tobacco and Arabidopsis TCP family were identified by genome-wide homologous alignment. The physicochemical properties, phylogenetic relationships and cis-acting elements were analyzed by bioinformatics. The homologous genes of AtTCP3/AtTCP4 were screened, and RT-qPCR was used to detect the changes of gene expression upon 20% PEG6000 treatment. The results show that tobacco contains 63 TCP family members. Their amino acid sequence length ranged from 89 aa to 596 aa, and their protein hydropathicity grand average of hydropathicity (GRAVY) ranged from -1.147 to 0.125. The isoelectric point (pI) ranges from 4.42 to 9.94, the number of introns is 0 to 3, and the subcellular location is all located in the nucleus. The results of conserved domain and phylogenetic relationship analysis showed that the tobacco TCP family can be divided into PCF, CIN and CYC/TB1 subfamilies, and each subfamily has a stable sequence. The results of cis-acting elements in gene promoter region showed that TCP family genes contain low docile acting elements (LTR) and a variety of stress and metabolic regulation related elements (MYB, MYC). Analysis of gene expression patterns showed that AtTCP3/AtTCP4 homologous genes (NtTCP6, NtTCP28, NtTCP30, NtTCP33, NtTCP42, NtTCP57, NtTCP63) accounted for 20% PEG6000 treatment significantly up-regulated/down-regulated expression, and NtTCP30 and NtTCP57 genes were selected as candidate genes in response to drought. The results of this study analyzed the TCP family in the tobacco genome and provided candidate genes for the study of drought-resistance gene function and variety breeding in tobacco.

Deciphering plant cell-cell communications using single-cell omics data.

Plants have various cell types that respond to different environmental factors, and cell-cell communication is the fundamental process that controls these plant responses. The emergence of single-cell techniques provides opportunities to explore features unique to each cell type and construct a comprehensive cell-cell communication (CCC) network. Although the most current successes of CCC inference were achieved in animal research, computational methods can also be directly applied to plants. This review describes the current major models for cell-cell communication inference and summarizes the computational tools based on single-cell omics datasets. In addition, we discuss the limitations of plant cell-cell communication research and propose new directions to expand the field in meaningful ways.