Plant Genome Analysis Research Articles

Doubling down on polyploid discoveries: Global advances in genomics and ecological impacts of polyploidy.

All flowering plants are now recognized as diploidized paleopolyploids (Jiao et al., 2011; One Thousand Plant Transcriptomes Initiative, 2019), and polyploid species comprise approximately 30% of contemporary plant species (Wood et al., 2009; Barker et al., 2016a). A major implication of these discoveries is that, to appreciate the evolution of plant diversity, we need to understand the fundamental biology of polyploids and diploidization. This need is broadly recognized by our community as there is a continued, growing interest in polyploidy as a research topic. Over the past 25 years, the sequencing and analysis of plant genomes has revolutionized our understanding of the importance of polyploid speciation to the evolution of land plants.

HaploMAGIC: accurate phasing and detection of recombination in multiparental populations despite genotyping errors.

Recombination is a key mechanism in breeding for promoting genetic variability. Multiparental populations (MPPs) constitute an excellent platform for precise genotype phasing, identification of genome-wide crossovers (COs), estimation of recombination frequencies, and construction of recombination maps. Here, we introduce haploMAGIC, a pipeline to detect COs in MPPs with single-nucleotide polymorphism (SNP) data by exploiting the pedigree relationships for accurate genotype phasing and inference of grandparental haplotypes. haploMAGIC applies filtering to prevent false-positive COs due to genotyping errors (GEs), a common problem in high-throughput SNP analysis of complex plant genomes. Hence, it discards haploblocks not reaching a specified minimum number of informative alleles. A performance analysis using populations simulated with AlphaSimR revealed that haploMAGIC improves upon existing methods of CO detection in terms of recall and precision, most notably when GE rates are high. Furthermore, we constructed recombination maps using haploMAGIC with high-resolution genotype data from 2 large multiparental populations of winter rapeseed (Brassica napus). The results demonstrate the applicability of the pipeline in real-world scenarios and showed good correlations in recombination frequency compared with alternative software. Therefore, we propose haploMAGIC as an accurate tool at CO detection with MPPs that shows robustness against GEs.

Genome sequencing provides potential strategies for drug discovery and synthesis

Medicinal plants are renowned for their abundant production of secondary metabolites, which exhibit notable pharmacological activities and great potential for drug development. The biosynthesis of secondary metabolites is highly intricate and influenced by various intrinsic and extrinsic factors, resulting in substantial species diversity and content variation. Consequently, precise regulation of secondary metabolite synthesis is of utmost importance. In recent years, genome sequencing has emerged as a valuable tool for investigating the synthesis and regulation of secondary metabolites in medicinal plants, facilitated by the widespread use of high-throughput sequencing technologies. This review highlights the latest advancements in genome sequencing within this field and presents several strategies for studying secondary metabolites. Specifically, the article elucidates how genome sequencing can unravel the pathways for secondary metabolite synthesis in medicinal plants, offering insights into the functions and regulatory mechanisms of participating enzymes. Comparative analyses of plant genomes allow identification of shared pathways of metabolite synthesis among species, thereby providing novel avenues for obtaining cost-effective biosynthetic intermediates. By examining individual genomic variations, genes or gene clusters associated with the synthesis of specific compounds can be discovered, indicating potential targets and directions for drug development and the exploration of alternative compound sources. Moreover, the advent of gene-editing technology has enabled the precise modifications of medicinal plant genomes. Optimization of specific secondary metabolite synthesis pathways becomes thus feasible, enabling the precise editing of target genes to regulate secondary metabolite production within cells. These findings serve as valuable references and lessons for future drug development endeavors, conservation of rare resources, and the exploration of new resources.

Application of modern biotechnology and bioinformatics approaches in agricultural sciences; A systematic review

In recent times, there has been a substantial surge in biological data owing to remarkable progress in molecular biology and genomic technology. With the growing demand for data storage and management tools, coupled with the surge in genetic data, there has been a corresponding rise in the availability of tools for analyzing, visualizing, modeling, and predicting large datasets. The proliferation of diverse datasets derived from a multitude of plant species has significantly broadened our comprehension, particularly within the realm of plant biotechnology. In the realm of "omics" methodology, proficient bioinformatics methodologies and tools are presently under development to facilitate expedited genome sequencing and the comprehensive analysis of plant genomes. The present investigation centers on the diverse applications of plant biotechnology bioinformatics and their potential to augment agricultural productivity. This study aims to analyze and assess the obstacles or constraints encountered in the field of plant biotechnology, which have impeded the advancement of plant genomes at a slower pace compared to animal genomes. To successfully examine the intricate details of the plant genome, bioinformatics methods must be strong egough which provides longer coverage. The area in plant biotechnology including the agricultural industry are expected to advance from the development of bioinformatics, as would mankind as a whole. The inclusion of computer based programs and data analysis tools in plant biotechnology has facilitated the indepth study of crop plants important for commerce and trade. Improved algorithm development is necessary to facilitate tasks such as data mining, analysis, comparison, and etc. Hence, the integration of novel techniques and information into bioinformatics for the advancement of plant biotechnology, the agricultural industry, and the future of humanity will heavily rely on the expertise of bioinformaticians.

Start codon targeted (SCoT) polymorphism marker in plant genome analysis: current status and prospects

The present review illustrates a comprehensive overview of the start codon targeted (SCoT) polymorphism marker and their utilization in various applications related to genetic and genomic studies. Start codon targeted (SCoT) polymorphism marker, a targeted fingerprinting marker technique, has gained considerable importance in plant genetics, genomics, and molecular breeding due to its many desirable features. SCoT marker targets the region flanking the start codon, a highly conserved region in plant genes. Therefore, it can distinguish genetic variations in a specific gene that link to a specific trait. It is a simple, novel, cost-effective, highly polymorphic, and reproducible molecular marker for which there is no need for prior sequence information. In the recent past, SCoT markers have been employed in many commercially important and underutilized plant species for a variety of applications, including genetic diversity analysis, interspecific/generic genetic relationships, cultivar/hybrid/species identification, sex determination, construction of linkage map, association mapping/analysis, differential gene expression, and genetic fidelity analysis of tissue culture-raised plants. The main aim of this review is to provide up-to-date information on SCoT markers and their application in many commercially important and underutilized plant species, mainly progress made in the last 8-10years.

Establishing MinION Sequencing and Genome Assembly Procedures for the Analysis of the Rooibos (Aspalathus linearis) Genome.

While plant genome analysis is gaining speed worldwide, few plant genomes have been sequenced and analyzed on the African continent. Yet, this information holds the potential to transform diverse industries as it unlocks medicinally and industrially relevant biosynthesis pathways for bioprospecting. Considering that South Africa is home to the highly diverse Cape Floristic Region, local establishment of methods for plant genome analysis is essential. Long-read sequencing is becoming standard procedure for plant genome research, as these reads can span repetitive regions of the DNA, substantially facilitating reassembly of a contiguous genome. With the MinION, Oxford Nanopore offers a cost-efficient sequencing method to generate long reads; however, DNA purification protocols must be adapted for each plant species to generate ultra-pure DNA, essential for these analyses. Here, we describe a cost-effective procedure for the extraction and purification of plant DNA and evaluate diverse genome assembly approaches for the reconstruction of the genome of rooibos (Aspalathus linearis), an endemic South African medicinal plant widely used for tea production. We discuss the pros and cons of nine tested assembly programs, specifically Redbean and NextDenovo, which generated the most contiguous assemblies, and Flye, which produced an assembly closest to the predicted genome size.

How do emerging long-read sequencing technologies function in transforming the plant pathology research landscape?

Long-read sequencing technologies are revolutionizing the sequencing and analysis of plant and pathogen genomes and transcriptomes, as well as contributing to emerging areas of interest in plant-pathogen interactions, disease management techniques, and the introduction of new plant varieties or cultivars. Long-read sequencing (LRS) technologies are progressively being implemented to study plants and pathogens of agricultural importance, which have substantial economic effects. The variability and complexity of the genome and transcriptome affect plant growth, development and pathogen responses. Overcoming the limitations of second-generation sequencing, LRS technology has significantly increased the length of a single contiguous read from a few hundred to millions of base pairs. Because of the longer read lengths, new analysis methods and tools have been developed for plant and pathogen genomics and transcriptomics. LRS technologies enable faster, more efficient, and high-throughput ultralong reads, allowing direct sequencing of genomes that would be impossible or difficult to investigate using short-read sequencing approaches. These benefits include genome assembly in repetitive areas, creating more comprehensive and exact genome determinations, assembling full-length transcripts, and detecting DNA and RNA alterations. Furthermore, these technologies allow for the identification of transcriptome diversity, significant structural variation analysis, and direct epigenetic mark detection in plant and pathogen genomic regions. LRS in plant pathology is found efficient for identifying and characterization of effectors in plants as well as known and unknown plant pathogens. In this review, we investigate how these technologies are transforming the landscape of determination and characterization of plant and pathogen genomes and transcriptomes efficiently and accurately. Moreover, we highlight potential areas of interest offered by LRS technologies for future study into plant-pathogen interactions, disease control strategies, and the development of new plant varieties or cultivars.

Plant growth-promoting effect and genomic analysis of the P. putida LWPZF isolated from C. japonicum rhizosphere

Plant growth-promoting rhizobacteria are a type of beneficial bacteria which inhabit in the rhizosphere and possess the abilities to promote plant growth. Pseudomonas putida LWPZF is a plant growth-promoting bacterium isolated from the rhizosphere soil of Cercidiphyllum japonicum. Inoculation treatment with LWPZF could significantly promote the growth of C. japonicum seedlings. P. putida LWPZF has a variety of plant growth-promoting properties, including the ability to solubilize phosphate, synthesize ACC deaminase and IAA. The P. putida LWPZF genome contained a circular chromosome (6,259,530 bp) and a circular plasmid (160,969 bp) with G+C contents of 61.75% and 58.25%, respectively. There were 5632 and 169 predicted protein-coding sequences (CDSs) on the chromosome and the plasmid respectively. Genome sequence analysis revealed lots of genes associated with biosynthesis of IAA, pyoverdine, ACC deaminase, trehalose, volatiles acetoin and 2,3-butanediol, 4-hydroxybenzoate, as well as gluconic acid contributing phosphate solubilization. Additionally, we identified many heavy metal resistance genes, including arsenate, copper, chromate, cobalt-zinc-cadmium, and mercury. These results suggest that P. putida LWPZF shows strong potential in the fields of biofertilizer, biocontrol and heavy metal contamination soil remediation. The data presented in this study will allow us to better understand the mechanisms of plant growth promotion, biocontrol, and anti-heavy metal of P. putida LWPZF.

Evaluation of methods for plant genomic DNA sequence analysis without DNA and PCR product purification

Genome-editing technologies are widely used to characterize gene functions and improve the features of agricultural plants. Although sequence analysis of gene editing target DNA is the most reliable method of screening gene-edited plants, the current DNA sequence analysis methods are time consuming and labor intensive because they include genomic DNA and polymerase chain reaction (PCR) product purification. In this study, seven methods were performed for sequence analysis of plant genomic DNA with and/or without genomic DNA and PCR product purification. Consequently, good-quality sequencing chromatograms were obtained using all methods. Results showed that the partial genomic DNA sequence of Nicotiana benthamiana and Arabidopsis thaliana could be sufficiently analyzed without plant genomic DNA and PCR product purification. Furthermore, screening of gene-edited N. benthamiana was successful using the present methods. Therefore, the tested methods could reduce the time, simplify the workflow of plant gene analysis, and help in screening gene-edited plants.

Genomic assessment of Stenotrophomonas indicatrix for improved sunflower plant.

Diverse agriculturally important microbes have been studied with known potential in plant growth promotion. Providing several opportunities, Stenotrophomonas species are characterized as promising plant enhancers, inducers, and protectors against environmental stressors. The S. indicatrix BOVIS40 isolated from the sunflower root endosphere possessed unique features, as genome insights into the Stenotrophomonas species isolated from oilseed crops in Southern Africa have not been reported. Plant growth-promotion screening and genome analysis of S. indicatrix BOVIS40 were presented in this study. The genomic information reveals various genes underlining plant growth promotion and resistance to environmental stressors. The genome of S. indicatrix BOVIS40 harbors genes involved in the degradation and biotransformation of organic molecules. Also, other genes involved in biofilm production, chemotaxis, and flagellation that facilitate bacterial colonization in the root endosphere and phytohormone genes that modulate root development and stress response in plants were detected in strain BOVIS40. IAA activity of the bacterial strain may be a factor responsible for root formation. A measurable approach to the S. indicatrix BOVIS40 lifestyle can strategically provide several opportunities in their use as bioinoculants in developing environmentally friendly agriculture sustainably. The findings presented here provide insights into the genomic functions of S. indicatrix BOVIS40, which has set a foundation for future comparative studies for a better understanding of the synergism among microbes inhabiting plant endosphere. Hence, highlighting the potential of S. indicatrix BOVIS40 upon inoculation under greenhouse experiment, thus suggesting its application in enhancing plant and soil health sustainably.

Impact of short-read sequencing on the misassembly of a plant genome

BackgroundAvailability of plant genome sequences has led to significant advances. However, with few exceptions, the great majority of existing genome assemblies are derived from short read sequencing technologies with highly uneven read coverages indicative of sequencing and assembly issues that could significantly impact any downstream analysis of plant genomes. In tomato for example, 0.6% (5.1 Mb) and 9.7% (79.6 Mb) of short-read based assembly had significantly higher and lower coverage compared to background, respectively.ResultsTo understand what the causes may be for such uneven coverage, we first established machine learning models capable of predicting genomic regions with variable coverages and found that high coverage regions tend to have higher simple sequence repeat and tandem gene densities compared to background regions. To determine if the high coverage regions were misassembled, we examined a recently available tomato long-read based assembly and found that 27.8% (1.41 Mb) of high coverage regions were potentially misassembled of duplicate sequences, compared to 1.4% in background regions. In addition, using a predictive model that can distinguish correctly and incorrectly assembled high coverage regions, we found that misassembled, high coverage regions tend to be flanked by simple sequence repeats, pseudogenes, and transposon elements.ConclusionsOur study provides insights on the causes of variable coverage regions and a quantitative assessment of factors contributing to plant genome misassembly when using short reads and the generality of these causes and factors should be tested further in other species.

Plants meet machines: Prospects in machine learning for plant biology

Plants meet machines: Prospects in machine learning for plant biology

Plant growth-promoting activities and genomic analysis of the stress-resistant Bacillus megaterium STB1, a bacterium of agricultural and biotechnological interest

In this work, the stress-resistant Bacillus megaterium STB1 is characterized and its ability to promote plant growth under normal and stress conditions is demonstrated. The genomic sequence of this bacterium, and a detailed analysis of the genes involved in facilitating its stress resistance and plant growth-promoting activities is also reported.The B. megaterium STB1 genome is rich in genetic elements involved in multiple stress resistance, xenobiotic degradation, pathogen antagonistic activities, and other traits related to soil and rhizosphere colonization. Moreover, genes participating in the biosynthesis of auxins and cytokinins, the modulation of polyamines, GABA, brassinosteroids and ethylene levels were also found.Ultimately, this study brings new insights into the role of B. megaterium as a plant growth-promoting bacterium and opens new opportunities for the development of novel strategies for agriculture and biotechnology.

The chromosome-scale reference genome of black pepper provides insight into piperine biosynthesis

Black pepper (Piper nigrum), dubbed the ‘King of Spices’ and ‘Black Gold’, is one of the most widely used spices. Here, we present its reference genome assembly by integrating PacBio, 10x Chromium, BioNano DLS optical mapping, and Hi-C mapping technologies. The 761.2 Mb sequences (45 scaffolds with an N50 of 29.8 Mb) are assembled into 26 pseudochromosomes. A phylogenomic analysis of representative plant genomes places magnoliids as sister to the monocots-eudicots clade and indicates that black pepper has diverged from the shared Laurales-Magnoliales lineage approximately 180 million years ago. Comparative genomic analyses reveal specific gene expansions in the glycosyltransferase, cytochrome P450, shikimate hydroxycinnamoyl transferase, lysine decarboxylase, and acyltransferase gene families. Comparative transcriptomic analyses disclose berry-specific upregulated expression in representative genes in each of these gene families. These data provide an evolutionary perspective and shed light on the metabolic processes relevant to the molecular basis of species-specific piperine biosynthesis.

Progress in plant genome sequencing: research directions

Since the first plant genome of Arabidopsis thaliana has been sequenced and published, genome sequencing technologies have undergone significant changes. New algorithms, sequencing technologies and bioinformatic approaches were adopted to obtain genome, transcriptome and exome sequences for model and crop species, which have permitted deep inferences into plant biology. As a result of an improved genome assembly and analysis methods, genome sequencing costs plummeted and the number of high-quality plant genome sequences is constantly growing. Consequently, more than 300 plant genome sequences have been published over the past twenty years. Although many of the published genomes are considered incomplete, they proved to be a valuable tool for identifying genes involved in the formation of economically valuable plant traits, for marker-assisted and genomic selection and for comparative analysis of plant genomes in order to determine the basic patterns of origin of various plant species. Since a high coverage and resolution of a genome sequence is not enough to detect all changes in complex samples, targeted sequencing, which consists in the isolation and sequencing of a specific region of the genome, has begun to develop. Targeted sequencing has a higher detection power (the ability to identify new differences/variants) and resolution (up to one basis). In addition, exome sequencing (the method of sequencing only protein-coding genes regions) is actively developed, which allows for the sequencing of non-expressed alleles and genes that cannot be found with RNA-seq. In this review, an analysis of sequencing technologies development and the construction of “reference” genomes of plants is performed. A comparison of the methods of targeted sequencing based on the use of the reference DNA sequence is accomplished.

Single-Cell Genomic Analysis in Plants

Individual cells in an organism are variable, which strongly impacts cellular processes. Advances in sequencing technologies have enabled single-cell genomic analysis to become widespread, addressing shortcomings of analyses conducted on populations of bulk cells. While the field of single-cell plant genomics is in its infancy, there is great potential to gain insights into cell lineage and functional cell types to help understand complex cellular interactions in plants. In this review, we discuss current approaches for single-cell plant genomic analysis, with a focus on single-cell isolation, DNA amplification, next-generation sequencing, and bioinformatics analysis. We outline the technical challenges of analysing material from a single plant cell, and then examine applications of single-cell genomics and the integration of this approach with genome editing. Finally, we indicate future directions we expect in the rapidly developing field of plant single-cell genomic analysis.

Plant Promoter Analysis: Identification and Characterization of Root Nodule Specific Promoter in the Common Bean.

The upstream sequences of gene coding sequences are termed as promoter sequences. Studying the expression patterns of promoters are very significant in understanding the gene regulation and spatiotemporal expression patterns of target genes. On the other hand, it is also critical to establish promoter evaluation tools and genetic transformation techniques that are fast, efficient, and reproducible. In this study, we investigated the spatiotemporal expression pattern of the rhizobial symbiosis-specific nodule inception (NIN) promoter of Phaseolus vulgaris in the transgenic hairy roots. Using plant genome databases and analysis tools we identified, isolated, and cloned the P. vulgaris NIN promoter in a transcriptional fusion to the chimeric reporter β-glucuronidase (GUS) GUS-enhanced::GFP. Further, this protocol describes a rapid and versatile system of genetic transformation in the P. vulgaris using Agrobacterium rhizogenes induced hairy roots. This system generates ≥2 cm hairy roots at 10 to 12 days after transformation. Next, we assessed the spatiotemporal expression of NIN promoter in Rhizobium inoculated hairy roots at periodic intervals of post-inoculation. Our results depicted by GUS activity show that the NIN promoter was active during the process of nodulation. Together, the present protocol demonstrates how to identify, isolate, clone, and characterize a plant promoter in the common bean hairy roots. Moreover, this protocol is easy to use in non-specialized laboratories.

DRDB: An Online Date Palm Genomic Resource Database.

Background: Date palm (Phoenix dactylifera L.) is a cultivated woody plant with agricultural and economic importance in many countries around the world. With the advantages of next generation sequencing technologies, genome sequences for many date palm cultivars have been released recently. Short sequence repeat (SSR) and single nucleotide polymorphism (SNP) can be identified from these genomic data, and have been proven to be very useful biomarkers in plant genome analysis and breeding.Results: Here, we first improved the date palm genome assembly using 130X of HiSeq data generated in our lab. Then 246,445 SSRs (214,901 SSRs and 31,544 compound SSRs) were annotated in this genome assembly; among the SSRs, mononucleotide SSRs (58.92%) were the most abundant, followed by di- (29.92%), tri- (8.14%), tetra- (2.47%), penta- (0.36%), and hexa-nucleotide SSRs (0.19%). The high-quality PCR primer pairs were designed for most (174,497; 70.81% out of total) SSRs. We also annotated 6,375,806 SNPs with raw read depth≥3 in 90% cultivars. To further reduce false positive SNPs, we only kept 5,572,650 (87.40% out of total) SNPs with at least 20% cultivars support for downstream analyses. The high-quality PCR primer pairs were also obtained for 4,177,778 (65.53%) SNPs. We reconstructed the phylogenetic relationships among the 62 cultivars using these variants and found that they can be divided into three clusters, namely North Africa, Egypt – Sudan, and Middle East – South Asian, with Egypt – Sudan being the admixture of North Africa and Middle East – South Asian cultivars; we further confirmed these clusters using principal component analysis. Moreover, 34,346 SSRs and 4,177,778 SNPs with PCR primers were assigned to shared cultivars for cultivar classification and diversity analysis. All these SSRs, SNPs and their classification are available in our database, and can be used for cultivar identification, comparison, and molecular breeding.Conclusion: DRDB is a comprehensive genomic resource database of date palm. It can serve as a bioinformatics platform for date palm genomics, genetics, and molecular breeding. DRDB is freely available at http://drdb.big.ac.cn/home.

Technical Review: Cytogenetic Tools for Studying Mitotic Chromosomes.

Significant advances in chromosome preparation and other techniques have greatly increased the potential of plant cytogenetics in recent years. Increase in longitudinal resolution using DNA extended fibers as well as new developments in imaging and signal amplification technologies have enhanced the ability of FISH to detect small gene targets. The combination of fluorescence in situ hybridization with immunocytochemistry allows the investigation of cell events, chromosomal rearrangements and chromatin features typical for plant nuclei. Chromosome manipulation techniques using microdissection and flow sorting have accelerated the analysis of complex plant genomes. Together, the different cytogenetic approaches are invaluable for the unravelling of detailed structures of plant chromosomes, which are of utmost importance for the study of genome properties, DNA replication and gene regulation. In this technical review, different cytogenetic approaches are discussed for the analysis of plant chromosomes, with a focus on mitotic chromosomes.

Application of molecular markers in plant genome analysis: a review

Advancement in the field of molecular biology has led to the development of various molecular markers which has revolutionized our understanding of the organization and evolution of plant genomes. Detection of genetic variation in plants offers an opportunity to understand the molecular basis of several biological phenomena. The reliability and efficiency of restriction digestion and polymerase chain reaction based random DNA markers have already proved their utility in taxonomical, evolutionary and ecological studies of plants. Progresses in the field of genomics and transcriptomics have enabled plant researchers to develop molecular markers derived from exon region of the genome which are termed as genic molecular markers (GMMs). GMMs are the part of the cDNA/EST sequences that mainly characterize the functional part of the genome. Next-generation DNA sequencing has also significantly contributed towards development of microRNA specific novel functional markers at the DNA level. This review focuses on the technical aspects of different molecular markers and their applications in the genome analysis.