Browsing by Author "Yan, Haidong"

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    Combinations of Small RNA, RNA, and Degradome Sequencing Uncovers the Expression Pattern of microRNA–mRNA Pairs Adapting to Drought Stress in Leaf and Root of Dactylis glomerata L.
    Ji, Yang; Chen, Peilin; Chen, Jing; Pennerman, Kayla K.; Liang, Xiaoyu; Yan, Haidong; Zhou, Sifan; Feng, Guangyan; Wang, Chengran; Yin, Guohua; Zhang, Xinquan; Hu, Yuanbin; Huang, Linkai (MDPI, 2018-10-11)
    Drought stress is a global problem, and the lack of water is a key factor that leads to agricultural shortages. MicroRNAs play a crucial role in the plant drought stress response; however, the microRNAs and their targets involved in drought response have not been well elucidated. In the present study, we used Illumina platform (https://www.illumina.com/) and combined data from miRNA, RNA, and degradome sequencing to explore the drought- and organ-specific miRNAs in orchardgrass (Dactylis glomerata L.) leaf and root. We aimed to find potential miRNA–mRNA regulation patterns responding to drought conditions. In total, 519 (486 conserved and 33 novel) miRNAs were identified, of which, 41 miRNAs had significant differential expression among the comparisons (p < 0.05). We also identified 55,366 unigenes by RNA-Seq, where 12,535 unigenes were differently expressed. Finally, our degradome analysis revealed that 5950 transcripts were targeted by 487 miRNAs. A correlation analysis identified that miRNA ata-miR164c-3p and its target heat shock protein family A (HSP70) member 5 gene comp59407_c0 (BIPE3) may be essential in organ-specific plant drought stress response and/or adaptation in orchardgrass. Additionally, Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) analyses found that “antigen processing and presentation” was the most enriched downregulated pathway in adaptation to drought conditions. Taken together, we explored the genes and miRNAs that may be involved in drought adaptation of orchardgrass and identified how they may be regulated. These results serve as a valuable genetic resource for future studies focusing on how plants adapted to drought conditions.
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    Comparative transcriptome study of switchgrass (Panicum virgatum L.) homologous autopolyploid and its parental amphidiploid responding to consistent drought stress
    Chen, Peilin; Chen, Jing; Sun, Min; Yan, Haidong; Feng, Guangyan; Wu, Bingchao; Zhang, Xinquan; Wang, Xiaoshan; Huang, Linkai (2020-10-15)
    Background Newly formed polyploids may experience short-term adaptative changes in their genome that may enhance the resistance of plants to stress. Considering the increasingly serious effects of drought on biofuel plants, whole genome duplication (WGD) may be an efficient way to proceed with drought resistant breeding. However, the molecular mechanism of drought response before/after WGD remains largely unclear. Results We found that autoploid switchgrass (Panicum virgatum L.) 8X Alamo had higher drought tolerance than its parent amphidiploid 4X Alamo using physiological tests. RNA and microRNA sequencing at different time points during drought were then conducted on 8X Alamo and 4X Alamo switchgrass. The specific differentially expressed transcripts (DETs) that related to drought stress (DS) in 8X Alamo were enriched in ribonucleoside and ribonucleotide binding, while the drought-related DETs in 4X Alamo were enriched in structural molecule activity. Ploidy-related DETs were primarily associated with signal transduction mechanisms. Weighted gene co-expression network analysis (WGCNA) detected three significant DS-related modules, and their DETs were primarily enriched in biosynthesis process and photosynthesis. A total of 26 differentially expressed microRNAs (DEmiRs) were detected, and among them, sbi-microRNA 399b was only expressed in 8X Alamo. The targets of microRNAs that were responded to polyploidization and drought stress all contained cytochrome P450 and superoxide dismutase genes. Conclusions This study explored the drought response of 8X and 4X Alamo switchgrass on both physiological and transcriptional levels, and provided experimental and sequencing data basis for a short-term adaptability study and drought-resistant biofuel plant breeding.
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    Development of tools to study the association of transposons to agronomic traits
    Yan, Haidong (Virginia Tech, 2020-05-21)
    Transposable elements (Transposons; TEs) constitute the majority of DNA in genomes and are a major source of genetic polymorphisms. TEs act as potential regulators of gene expression and lead to phenotypic plasticity in plants and animals. In crops, several TEs were identified to influence alleles associated with important agronomic traits, such as apical dominance in maize and seed number in rice. Crops may harbor more TE-mediated genetic regulations than expected in view of multifunctional TEs in genomes. However, tools that accurately annotate TEs and clarify their associations with agronomic traits are still lacking, which largely limits applications of TEs in crop breeding. Here we 1) evaluate performances of popular tools and strategies to identify TEs in genomes, 2) develop a tool 'DeepTE' to annotate TEs based on deep learning models, and 3) develop a tool 'TE-marker' to identify potential TE-regulated alleles associated with agronomic traits. As a result, we propose a series of recommendations and a guideline to develop a comprehensive library to precisely identify TEs in genomes. Secondly, 'DeepTE' classifies TEs into 15-24 super families according to sequences from plants, metazoans, and fungi. For unknown sequences, this tool can distinguish non-TEs and TEs in plant species. Finally, the 'TE-marker' tool builds a TE-based marker system that is able to cluster rice populations similar to a classical SNP marker approach. This system can also detect association peaks that are equivalent to the ones produced by SNP markers. 'TE-marker' is a novel complementary approach to the classical SNP markers that it assists in revealing population structures and in identifying alleles associated with agronomic traits.
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    Exploring transposable element-based markers to identify allelic variations underlying agronomic traits in rice
    Yan, Haidong; Haak, David C.; Li, Song; Huang, Linkai; Bombarely, Aureliano (Elsevier, 2022-05-09)
    Transposable elements (TEs) are a major force in the production of new alleles during domestication; nevertheless, their use in association studies has been limited because of their complexity. We have developed a TE genotyping pipeline (TEmarker) and applied it to whole-genome genome-wide association study (GWAS) data from 176 Oryza sativa subsp. japonica accessions to identify genetic elements associated with specific agronomic traits. TE markers recovered a large proportion (69%) of single-nucleotide polymorphism (SNP)-based GWAS peaks, and these TE peaks retained ca. 25% of the SNPs. The use of TEs in GWASs may reduce false positives associated with linkage disequilibrium (LD) among SNP markers. A genome scan revealed positive selection on TEs associated with agronomic traits. We found several cases of insertion and deletion variants that potentially resulted from the direct action of TEs, including an allele of LOC_Os11g08410 associated with plant height and panicle length traits. Together, these findings reveal the utility of TE markers for connecting genotype to phenotype and suggest a potential role for TEs in influencing phenotypic variations in rice that impact agronomic traits.
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    Genome assembly provides insights into the genome evolution and flowering regulation of orchardgrass
    Huang, Linkai; Feng, Guangyan; Yan, Haidong; Zhang, Zhongren; Bushman, Bradley Shaun; Wang, Jianping; Bombarely, Aureliano; Li, Mingzhou; Yang, Zhongfu; Nie, Gang; Xie, Wengang; Xu, Lei; Chen, Peilin; Zhao, Xinxin; Jiang, Wenkai; Zhang, Xinquan (2020-02)
    Orchardgrass (Dactylis glomerata L.) is an important forage grass for cultivating livestock worldwide. Here, we report an similar to 1.84-Gb chromosome-scale diploid genome assembly of orchardgrass, with a contig N50 of 0.93 Mb, a scaffold N50 of 6.08 Mb and a super-scaffold N50 of 252.52 Mb, which is the first chromosome-scale assembled genome of a cool-season forage grass. The genome includes 40 088 protein-coding genes, and 69% of the assembled sequences are transposable elements, with long terminal repeats (LTRs) being the most abundant. The LTRretrotransposons may have been activated and expanded in the grass genome in response to environmental changes during the Pleistocene between 0 and 1 million years ago. Phylogenetic analysis reveals that orchardgrass diverged after rice but before three Triticeae species, and evolutionarily conserved chromosomes were detected by analysing ancient chromosome rearrangements in these grass species. We also resequenced the whole genome of 76 orchardgrass accessions and found that germplasm from Northern Europe and East Asia clustered together, likely due to the exchange of plants along the 'Silk Road' or other ancient trade routes connecting the East and West. Last, a combined transcriptome, quantitative genetic and bulk segregant analysis provided insights into the genetic network regulating flowering time in orchardgrass and revealed four main candidate genes controlling this trait. This chromosome-scale genome and the online database of orchardgrass developed here will facilitate the discovery of genes controlling agronomically important traits, stimulate genetic improvement of and functional genetic research on orchardgrass and provide comparative genetic resources for other forage grasses.
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    Integration of small RNAs and transcriptome sequencing uncovers a complex regulatory network during vernalization and heading stages of orchardgrass (Dactylis glomerata L.)
    Feng, Guangyan; Xu, Lei; Wang, Jianping; Nie, Gang; Bushman, Bradley Shaun; Xie, Wengang; Yan, Haidong; Yang, Zhongfu; Guan, Hao; Huang, Linkai; Zhang, Xinquan (2018-10-03)
    Background Flowering is a critical reproductive process in higher plants. Timing of optimal flowering depends upon the coordination among seasonal environmental cues. For cool season grasses, such as Dactylis glomerata, vernalization induced by low temperature provides competence to initiate flowering after prolonged cold. We combined analyses of the transcriptome and microRNAs (miRNAs) to generate a comprehensive resource for regulatory miRNAs and their target circuits during vernalization and heading stages. Results A total of 3,846 differentially expressed genes (DEGs) and 69 differentially expressed miRNAs were identified across five flowering stages. The expression of miR395, miR530, miR167, miR396, miR528, novel_42, novel_72, novel_107, and novel_123 demonstrated significant variations during vernalization. These miRNA targeted genes were involved in phytohormones, transmembrane transport, and plant morphogenesis in response to vernalization. The expression patterns of DEGs related to plant hormones, stress responses, energy metabolism, and signal transduction changed significantly in the transition from vegetative to reproductive phases. Conclusions Five hub genes, c136110_g1 (BRI1), c131375_g1 (BZR1), c133350_g1 (VRN1), c139830_g1 (VIN3), and c125792_g2 (FT), might play central roles in vernalization response. Our comprehensive analyses have provided a useful platform for investigating consecutive transcriptional and post-transcriptional regulation of critical phases in D. glomerata and provided insights into the genetic engineering of flowering-control in cereal crops.
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    De novo Transcriptome Analysis and Molecular Marker Development of Two Hemarthria Species
    Huang, Xiu; Yan, Haidong; Zhang, Xinquan; Zhang, Jian; Frazier, Taylor P.; Huang, Dejun; Lu, Lu; Huang, Linkai; Liu, Wei; Peng, Yan; Ma, Xiao; Yan, Yan-Hong (Frontiers, 2016-04-18)
    Hemarthria R. Br. is an important genus of perennial forage grasses that is widely used in subtropical and tropical regions. Hemarthria grasses have made remarkable contributions to the development of animal husbandry and agro-ecosystem maintenance; however, there is currently a lack of comprehensive genomic data available for these species. In this study, we used Illumina high-throughput deep sequencing to characterize of two agriculturally important Hemarthria materials, H. compressa "Yaan" and H. altissima "1110." Sequencing runs that used each of four normalized RNA samples from the leaves or roots of the two materials yielded more than 24 million high-quality reads. After de novo assembly, 137,142 and 77,150 unigenes were obtained for "Yaan" and "1110," respectively. In addition, a total of 86,731 "Yawn" and 48,645 "1110" unigenes were successfully annotated. After consolidating the unigenes for both materials, 42,646 high-quality SNPs were identified in 10,880 unigenes and 10,888 SSRs were identified in 8330 unigenes. To validate the identified markers, high quality PCR primers were designed for both SNPs and SSRs. We randomly tested 16 of the SNP primers and 54 of the SSR primers and found that the majority of these primers successfully amplified the desired PCR product. In addition, high cross-species transferability (61.11-87.04%) of SSR markers was achieved for four other Poaceae species. The amount of RNA sequencing data that was generated for these two Hemarthria species greatly increases the amount of genomic information available for Hemarthria and the SSR and SNP markers identified in this study will facilitate further advancements in genetic and molecular studies of the Hemarthria genus.
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    Pangenomic analysis identifies structural variation associated with heat tolerance in pearl millet
    Yan, Haidong; Sun, Min; Zhang, Zhongren; Jin, Yarong; Zhang, Ailing; Lin, Chuang; Wu, Bingchao; He, Min; Xu, Bin; Wang, Jing; Qin, Peng; Mendieta, John Pablo; Nie, Gang; Wang, Jianping; Jones, Chris S. S.; Feng, Guangyan; Srivastava, Rakesh K. K.; Zhang, Xinquan; Bombarely, Aureliano; Luo, Dan; Jin, Long; Peng, Yuanying; Wang, Xiaoshan; Ji, Yang; Tian, Shilin; Huang, Linkai (Nature Portfolio, 2023-03)
    Pearl millet is an important cereal crop worldwide and shows superior heat tolerance. Here, we developed a graph-based pan-genome by assembling ten chromosomal genomes with one existing assembly adapted to different climates worldwide and captured 424,085 genomic structural variations (SVs). Comparative genomics and transcriptomics analyses revealed the expansion of the RWP-RK transcription factor family and the involvement of endoplasmic reticulum (ER)-related genes in heat tolerance. The overexpression of one RWP-RK gene led to enhanced plant heat tolerance and transactivated ER-related genes quickly, supporting the important roles of RWP-RK transcription factors and ER system in heat tolerance. Furthermore, we found that some SVs affected the gene expression associated with heat tolerance and SVs surrounding ER-related genes shaped adaptation to heat tolerance during domestication in the population. Our study provides a comprehensive genomic resource revealing insights into heat tolerance and laying a foundation for generating more robust crops under the changing climate. A graph-based pan-genome constructed using de novo genome assemblies of ten pearl millet accessions adapted to different climates worldwide identifies structural variations and their contribution to heat tolerance in pearl millet.
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    Reference Gene Selection for Quantitative Real-Time Reverse-Transcriptase PCR in Annual Ryegrass (Lolium multiflorum) Subjected to Various Abiotic Stresses
    Liu, Qiuxu; Qi, Xiao; Yan, Haidong; Huang, Linkai; Nie, Gang; Zhang, Xinquan (MDPI, 2018-01-16)
    To select the most stable reference genes in annual ryegrass (Lolium multiflorum), we studied annual ryegrass leaf tissues exposed to various abiotic stresses by qRT-PCR and selected 11 candidate reference genes, i.e., 18S rRNA, E2, GAPDH, eIF4A, HIS3, SAMDC, TBP-1, Unigene71, Unigene77, Unigene755, and Unigene14912. We then used GeNorm, NormFinder, and BestKeeper to analyze the expression stability of these 11 genes, and used RefFinder to comprehensively rank genes according to stability. Under different stress conditions, the most suitable reference genes for studies of leaf tissues of annual ryegrass were different. The expression of the eIF4A gene was the most stable under drought stress. Under saline-alkali stress, Unigene14912 has the highest expression stability. Under acidic aluminum stress, SAMDC expression stability was highest. Under heavy metal stress, Unigene71 expression had the highest stability. According to the software analyses, Unigene14912, HIS3, and eIF4A were the most suitable for analyses of abiotic stress in tissues of annual ryegrass. GAPDH was the least suitable reference gene. In conclusion, selecting appropriate reference genes under abiotic stress not only improves the accuracy of annual ryegrass gene expression analyses, but also provides a theoretical reference for the development of reference genes in plants of the genus Lolium.
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    siRNAs regulate DNA methylation and interfere with gene and lncRNA expression in the heterozygous polyploid switchgrass
    Yan, Haidong; Bombarely, Aureliano; Xu, Bin; Frazier, Taylor P.; Wang, Chengran; Chen, Peilin; Chen, Jing; Hasing, Tomas; Cui, Chenming; Zhang, Xinquan; Zhao, Bingyu Y.; Huang, Linkai (2018-07-24)
    Background Understanding the DNA methylome and its relationship with non-coding RNAs, including microRNAs (miRNAs) and long non-coding RNAs (lncRNAs), is essential for elucidating the molecular mechanisms underlying key biological processes in plants. Few studies have examined the functional roles of the DNA methylome in grass species with highly heterozygous polyploid genomes. Results We performed genome-wide DNA methylation profiling in the tetraploid switchgrass (Panicum virgatum L.) cultivar ‘Alamo’ using bisulfite sequencing. Single-base-resolution methylation patterns were observed in switchgrass leaf and root tissues, which allowed for characterization of the relationship between DNA methylation and mRNA, miRNA, and lncRNA populations. The results of this study revealed that siRNAs positively regulate DNA methylation of the mCHH sites surrounding genes, and that DNA methylation interferes with gene and lncRNA expression in switchgrass. Ninety-six genes covered by differentially methylated regions (DMRs) were annotated by GO analysis as being involved in stimulus-related processes. Functionally, 82% (79/96) of these genes were found to be hypomethylated in switchgrass root tissue. Sequencing analysis of lncRNAs identified two lncRNAs that are potential precursors of miRNAs, which are predicted to target genes that function in cellulose biosynthesis, stress regulation, and stem and root development. Conclusions This study characterized the DNA methylome in switchgrass and elucidated its relevance to gene and non-coding RNAs. These results provide valuable genomic resources and references that will aid further epigenetic research in this important biofuel crop.
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    The ‘Tommy Atkins’ mango genome reveals candidate genes for fruit quality
    Bally, Ian S. E.; Bombarely, Aureliano; Chambers, Alan H.; Cohen, Yuval; Dillon, Natalie L.; Innes, David J.; Islas-Osuna, María A.; Kuhn, David N.; Mueller, Lukas A.; Ophir, Ron; Rambani, Aditi; Sherman, Amir; Yan, Haidong (2021-02-22)
    Background Mango, Mangifera indica L., an important tropical fruit crop, is grown for its sweet and aromatic fruits. Past improvement of this species has predominantly relied on chance seedlings derived from over 1000 cultivars in the Indian sub-continent with a large variation for fruit size, yield, biotic and abiotic stress resistance, and fruit quality among other traits. Historically, mango has been an orphan crop with very limited molecular information. Only recently have molecular and genomics-based analyses enabled the creation of linkage maps, transcriptomes, and diversity analysis of large collections. Additionally, the combined analysis of genomic and phenotypic information is poised to improve mango breeding efficiency. Results This study sequenced, de novo assembled, analyzed, and annotated the genome of the monoembryonic mango cultivar ‘Tommy Atkins’. The draft genome sequence was generated using NRGene de-novo Magic on high molecular weight DNA of ‘Tommy Atkins’, supplemented by 10X Genomics long read sequencing to improve the initial assembly. A hybrid population between ‘Tommy Atkins’ x ‘Kensington Pride’ was used to generate phased haplotype chromosomes and a highly resolved phased SNP map. The final ‘Tommy Atkins’ genome assembly was a consensus sequence that included 20 pseudomolecules representing the 20 chromosomes of mango and included ~ 86% of the ~ 439 Mb haploid mango genome. Skim sequencing identified ~ 3.3 M SNPs using the ‘Tommy Atkins’ x ‘Kensington Pride’ mapping population. Repeat masking identified 26,616 genes with a median length of 3348 bp. A whole genome duplication analysis revealed an ancestral 65 MYA polyploidization event shared with Anacardium occidentale. Two regions, one on LG4 and one on LG7 containing 28 candidate genes, were associated with the commercially important fruit size characteristic in the mapping population. Conclusions The availability of the complete ‘Tommy Atkins’ mango genome will aid global initiatives to study mango genetics.
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    Transcriptional Changes in Pearl Millet Leaves under Heat Stress
    Huang, Dejun; Sun, Min; Zhang, Ailing; Chen, Jishan; Zhang, Jian; Lin, Chuang; Zhang, Huan; Lu, Xiaowen; Wang, Xiaoshan; Yan, Haidong; Tang, Jianan; Huang, Linkai (MDPI, 2021-10-28)
    High-temperature stress negatively affects the growth and development of plants, and therefore threatens global agricultural safety. Cultivating stress-tolerant plants is the current objective of plant breeding programs. Pearl millet is a multi-purpose plant, commonly used as a forage but also an important food staple. This crop is very heat-resistant and has a higher net assimilation rate than corn under high-temperature stress. However, the response of heat resistant pearl millet has so far not been studied at the transcriptional level. In this study, transcriptome sequencing of pearl millet leaves exposed to different lengths of heat treatment (1 h, 48 h and 96 h) was conducted in order to investigate the molecular mechanisms of the heat stress response and to identify key genes related to heat stress. The results showed that the amount of heat stress-induced DEGs in leaves differs with the length of exposure to high temperatures. The highest value of DEGs (8286) was observed for the group exposed to heat stress for 96 h, while the other two treatments showed lower DEGs values of 4659 DEGs after 1 h exposure and 3981 DEGs after 48 h exposure to heat stress. The DEGs were mainly synthesized in protein folding pathways under high-temperature stress after 1 h exposure. Moreover, a large number of genes encoding ROS scavenging enzymes were activated under heat stress for 1 h and 48 h treatments. The flavonoid synthesis pathway of pearl millet was enriched after heat stress for 96 h. This study analyzed the transcription dynamics under short to long-term heat stress to provide a theoretical basis for the heat resistance response of pearl millet.
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    Transcriptome analysis of heat stress and drought stress in pearl millet based on Pacbio full-length transcriptome sequencing
    Sun, Min; Huang, Dejun; Zhang, Ailing; Khan, Imran; Yan, Haidong; Wang, Xiaoshan; Zhang, Xinquan; Zhang, Jian; Huang, Linkai (2020-07-08)
    Background Heat and drought are serious threats for crop growth and development. As the sixth largest cereal crop in the world, pearl millet can not only be used for food and forage but also as a source of bioenergy. Pearl millet is highly tolerant to heat and drought. Given this, it is considered an ideal crop to study plant stress tolerance and can be used to identify heat-resistant genes. Results In this study, we used Pacbio sequencing data as a reference sequence to analyze the Illumina data of pearl millet that had been subjected to heat and drought stress for 48 h. By summarizing previous studies, we found 26,299 new genes and 63,090 new transcripts, and the number of gene annotations increased by 20.18%. We identified 2792 transcription factors and 1223 transcriptional regulators. There were 318 TFs and 149 TRs differentially expressed under heat stress, and 315 TFs and 128 TRs were differentially expressed under drought stress. We used RNA sequencing to identify 6920 genes and 6484 genes differentially expressed under heat stress and drought stress, respectively. Conclusions Through Pacbio sequencing, we have identified more new genes and new transcripts. On the other hand, comparing the differentially expressed genes under heat tolerance with the DEGs under drought stress, we found that even in the same pathway, pearl millet responds with a different protein.
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    Whole-Genome Resequencing of Seven Eggplant (Solanum melongena) and One Wild Relative (S. incanum) Accessions Provides New Insights and Breeding Tools for Eggplant Enhancement
    Gramazio, Pietro; Yan, Haidong; Hasing, Tomas; Vilanova, Santiago; Prohens, Jaime; Bombarely, Aureliano (2019-10-08)
    Whole-genome resequencing provides information of great relevance for crop genetics, evolution, and breeding. Here, we present the first whole-genome resequencing study using seven eggplant (Solanum melongena) and one wild relative (Solanum incanum) accessions. These eight accessions were selected for displaying a high phenotypic and genetic diversity and for being the founder parents of an eggplant multiparent advanced generation intercrosses population. By resequencing at an average depth of 19.8x and comparing to the high-quality reference genome "67/3" over 10 million high-reliable polymorphisms were discovered, of which over 9 million (84.5%) were single nucleotide polymorphisms and more than 700,000 (6.5%) InDels. However, while for the S. melongena accessions, the variants identified ranged from 0.8 to 1.3 million, over 9 million were detected for the wild S. incanum. This confirms the narrow genetic diversity of the domesticated eggplant and suggests that introgression breeding using wild relatives can efficiently contribute to broadening the genetic basis of this crop. Differences were observed among accessions for the enrichment in genes regulating important biological processes. By analyzing the distribution of the variants, we identified the potential footprints of old introgressions from wild relatives that can help to unravel the unclear domestication and breeding history. The comprehensive annotation of these eight genomes and the information provided in this study represents a landmark in eggplant genomics and allows the development of tools for eggplant genetics and breeding.