Development Of Molecular Markers Research Articles

Comparative Analyses of Acacia Plastomes to Detect Mutational Hotspots and Barcode Sites for the Identification of Important Timber Species

The genus Acacia is a crucial source of industrial papermaking and timber, possessing significant economic value. Due to their nitrogen-fixing ability, rapid growth, and high tolerance to stress, Acacia species have become integral to short-term industrial timber forests, particularly in southern China and various other regions worldwide. Nevertheless, comparative genomic analyses of Acacia species remain limited, impeding the development of molecular markers for identifying and distinguishing between distinct Acacia species as well as distinguishing their wood counterparts from other tree species. To address this gap, we generated plastomic sequences from eight species commonly cultivated in Chinese plantation forests and compared them with existing data. Our findings revealed a generally conserved structure and gene content within the plastid genomes of Acacia. Notably, the dN/dS ratio of genes from distinct functional groups varied, particularly between ribosomal proteins and photosynthesis genes. Furthermore, phylogenetic analyses and sequence comparisons suggest that the introduction of Acacia to China may have been less diverse than previously thought or that interspecies introgression has occurred more frequently than previously documented. This study offers a valuable starting point for future research in this area and for improving timber quality through marker-assisted breeding.

Analyzing genetic diversity of chloroplast genomes in Liliales

AbstractLiliales is a monocotyledonous order and contains both photosynthetic and mycoheterotrophic species that distribute locally or worldwide. In this study, thegenetic diversity of chloroplast genomes in Liliales was explored regarding theirnucleotide diversity and repeated composition. The analysis of nucleotide diversity revealed various hotspots in large and small single-copy regions whereas the IR regions had low sequence divergence. Although each family has specific hotspots, the rps15-ycf1 region was commonly found as a highly variable area in the cpDNA of observed taxa. In the cpDNA of Liliales, mononucleotide simple sequence repeat (SSR) is the most common type. The majority of SSRs are located in non-coding regions. Similarly, more long repeats were found in non-coding areas than in coding sequences. Additionally, the complement repeat exceeds forward type in the cpDNA of Liliales. The highest number of long repeats was found in Corsia dispar whereas that of SSRswas detected in Smilax china. The results of nucleotide diversity and repeat analyses provided fundamental information for further studies on population genetics, molecular marker development and evolutionary history of Liliales.® 2022 Journal of Science and Technology - NTTU

Development and validation of genome-wide SSR molecular markers of Tapes dorsatus.

Tapes dorsatus is an economically important benthic animal in the Beibu Gulf of China. However, the deficiency of microsatellite markers has hindered the study of its genetics. The development of microsatellite markers will provide useful tools for genetic improvement, variety identification, phylogenetic analysis and resource conservation. Within the genome sequence, 145,008 simple sequence repeats (SSRs) were identified, and 29,691 primer pairs were designed successfully. A total of 100 primer pairs were randomly synthesized for testing, and 93 primers yielded products. Sixty highly polymorphic primers were used to reveal the genetic diversity of 50 T. dorsatus individuals. The average number of alleles (Na) of the population was 10.40; the average number of effective alleleswas 6.16, the average expected heterozygosity (He) was 0.82, and the average polymorphic information contentwas 0.80. The genetic structure of the population was detected, by which the population could be divided into three subpopulations. We identified 145,008 SSRs in the genome of T. dorsatus and designed 29,691 primer pairs in this study. Of 100 synthesized primers, 60 were highly polymorphic and used to reveal the genetic diversity and structure of the population. The SSR markers identified here will provide useful tools and a foundation for genetic diversity, linkage mapping and molecular marker-aided breeding in T. dorsatus.

Development of High-Throughput Molecular Markers for Soybean Breeding

Development of High-Throughput Molecular Markers for Soybean Breeding

Comparative analysis of chloroplast genome and evolutionary history of Hemerocallis.

Members of the genus Hemerocallis have significant value as ornamental, edible, and medicinal plants, particularly in China, where they have been utilized for thousands of years as both a vegetable and Traditional Chinese Medicine. Hemerocallis species exhibit strict control over flowering time, with individuals flowering either diurnally or nocturnally. However, our understanding of the evolutionary history of this genus, especially concerning important horticultural traits, remains limited. In this study, sequencing and assembly efforts were conducted on 73 samples within the Hemerocallis genus. All accessions were classified into two distinct groups based on their diurnal (daylilies) or nocturnal (nightlilies) flowering habits. Comparative analysis of the chloroplast genomes from these two groups identified fifteen variant hotspot regions, including fourteen SNPs and one deletion, which hold promise for the development of molecular markers for interspecific identification. Phylogenetic trees, generated through both maximum-likelihood and Bayesian inference methods using 76 shared protein-coding sequences, revealed that diurnal flowering evolved prior to nocturnal flowering. The divergence between the two groups is estimated to have occurred approximately 0.82 MYA (95% CI: 0.35-1.45 MYA). The ancestral state of Hemerocallis is hypothesized to have featured diurnal flowering with orange yellow petals. This study marks the first reconstruction of the evolutionary history and ancestral state of the genus Hemerocallis. The findings contribute significantly to our understanding of the adaptation and speciation history within the genus.

Cabbage: Breeding and Genomics

Plant breeding has had a tremendous influence on crop improvement. However, due to dwindling germplasm resources, identification of variability for incorporation into new cultivars is becoming more difficult. Therefore, there has been recourse to alternative approaches including mutagenesis, tissue culture and genetic transformation to aid breeding programs. Development and application of molecular markers derived from genes, commonly called genic markers or sometimes functional markers, is gaining momentum in plant genetics and breeding. Presently, marker discovery, genotyping and molecular breeding practices would be routine in crop improvement in many crop species including Brassica. Molecular markers are known as particularly effective and reliable tools for the characterization of genome architectures and the investigation of gene polymorphisms in cabbage also. Cabbage breeding, genomics and their applications have been discussed in detailed in this review article.

Chili: Breeding and Genomics

Chili or hot pepper is an important vegetable cum spice crop, which has huge commercial applications in food, nutrition, pharmaceutical and cosmetic industries. The Capsicum genus originated in Central America with about 35 species, of which five are domesticated for cultivation. Color and pungency-related components such as capsanthins, capsaicinoids and capsinoids are unique to Capsicum genus. There exists large variability in the genus and genetic resources have been explored for breeding varieties/ hybrids with resistance/ tolerance to biotic and abiotic stresses with enhanced fruit quality. In the present climate change scenario, there is an evolution of various pests and diseases, which requires continuous exploration of germplasm including related cultivated and wild species for novel genes/alleles. Conventional breeding has led to the development of improved lines/varieties/hybrids. Whole genome sequence information and the pan genome of chili are publicly available, which can help in the development of molecular markers associated with key traits of importance. Advancements in genomic resources and next-generation sequencing techniques can be employed to accelerate breeding programs.

Development of a ferroptosis-based molecular markers for predicting RFS in prostate cancer patients

The goal of this study was to develop a ferroptosis-based molecular signature that can predict recurrence-free survival (RFS) in patients with prostate cancer (PCa). In this study, we obtained ferroptosis-related genes (FRGs) in FerrDb database and clinical transcriptome data in TCGA database and GEO database. Consensus cluster analysis was used to identify three molecular markers of ferroptosis in PCa with differential expression of 40 FRGs, including PD-L1 expression levels. We conducted a new ferroptosis-related signature for PCa RFS using four FRGs identified through univariate and multivariate Cox regression analyses. The signature was validated in the training, testing, and validation cohorts, and it demonstrated remarkable results in the area under the time-dependent receiver operating characteristic (ROC) curve of 0.757, 0.715, and 0.732, respectively. Additionally, we observed that younger patients, those with stage T III and stage T IV, stage N0, cluster 1, and cluster 2 PCa were more accurately predicted by the signature as independent predictors of RFS. DU-145 and RWPE-1 cells were successfully analyzed by qRT-PCR and Western blot for ASNS, GPT2, RRM2, and NFE2L2. In summary, we developed a novel ferroptosis-based signature for RFS in PC, utilizing four FRGs identified through univariate and multivariate Cox regression analyses. This signature was rigorously validated across training, testing, and validation cohorts, demonstrating exceptional performance as evidenced by its ROC curves. Notably, our findings indicate that this signature is particularly effective as an independent predictor of RFS in younger patients or those with stage T III and T IV, stage N0, and in clusters 1 and 2. Finally, we confirmed the expression of these four FRGs in DU-145 and RWPE-1 cell lines.

Genetic mapping of quantitative trait loci controlling smut resistance in Louisiana sugarcane using a bi-parental mapping population

Sugarcane smut, caused by Sporisorium scitamineum, is a major disease worldwide. Breeding resistant cultivars is the main management strategy. However, occasional failure to detect susceptible clones due to unfavorable environmental conditions, inconsistency in disease ratings between experiments, and continued clone losses due to pathogen adaptability are some of the challenges for this strategy. The development and use of molecular markers associated with smut resistance may overcome these limitations allowing more accurate identification of resistant parents and progeny. A genetic analysis was conducted using an inoculated population of 162 F1 progeny from a biparental cross between susceptible/female and resistant/male parents to identify quantitative trait loci (QTLs) associated with resistance. A total of 1574 single dose (SD) single-nucleotide polymorphisms (SNP) markers used to construct a genetic map resulted in 253 linkage groups (LGs) of which 150 LGs were assigned to the female parent and 204 LG to the male parent with a genome coverage of 24,580 cM. (Composite) interval mapping with selective sub-populations identified six consistent QTLs that cumulatively explained 25.74% of the phenotypic variance with LOD scores ranging from 3.17 to 23.7. Four out of 12 SNP markers closest to the QTL peaks had effect >10 for resistance in the heterozygous condition. Genes known to be involved in disease resistance, such as cellulose synthase, expansin, protein degradation, and receptor-kinase were linked to the genomic regions associated with smut resistance. The markers upon validation in different populations can be utilized for marker-assisted selection.

Genomic Insights into Molecular Regulation Mechanisms of Intramuscular Fat Deposition in Chicken.

Intramuscular fat (IMF) plays an important role in the tenderness, water-holding capacity, and flavor of chicken meat, which directly affect meat quality. In recent years, regulatory mechanisms underlying IMF deposition and the development of effective molecular markers have been hot topics in poultry genetic breeding. Therefore, this review focuses on the current understanding of regulatory mechanisms underlying IMF deposition in chickens, which were identified by multiple genomic approaches, including genome-wide association studies, whole transcriptome sequencing, proteome sequencing, single-cell RNA sequencing (scRNA-seq), high-throughput chromosome conformation capture (HiC), DNA methylation sequencing, and m6A methylation sequencing. This review comprehensively and systematically describes genetic and epigenetic factors associated with IMF deposition, which provides a fundamental resource for biomarkers of IMF deposition and provides promising applications for genetic improvement of meat quality in chicken.

Development of novel specific molecular markers for the Sw-5b gene to assist with tomato spotted wilt virus-resistant tomato breeding

Tomato spotted wilt virus (TSWV) is a plant pathogen that causes devastating tomato yield losses worldwide. The Sw-5b gene is one of the most effective resistance genes for TSWV control in tomato plants, and has been widely used in resistance breeding. Molecular markers are specific DNA sequences with known locations on the chromosome; they are indispensable tools in marker-assisted selection, which detects the presence of target genes to expedite breeding. We developed gene-specific molecular markers for Sw-5b to facilitate the accurate distinction of resistance (Sw-5bR) and susceptibility (Sw-5bS) alleles of Sw-5b. Using these markers, we successfully detected Sw-5b and determined its genotype (homozygous Sw-5bR, heterozygous Sw-5bR/S, or homozygous Sw-5bS) in six tomato varieties. Then we successfully applied these markers to 46 commercial tomato cultivars to detect and determine the genotype of Sw-5b. The results revealed a striking absence of the Sw-5bR gene and high TSWV susceptibility among most of the analyzed commercial cultivars. With the assistance of the novel Sw-5b-specific molecular markers, we generated a TSWV-resistant and homozygous Sw-5bR Micro-Tom tomato line, demonstrating the practical application of these markers in plant breeding. In summary, we developed novel gene-specific molecular markers for Sw-5b, and applied them to distinguish Sw-5b alleles for TSWV resistance or susceptibility. This marker set provides a valuable tool for breeding TSWV-resistant tomato varieties.

Comparative Analysis of Six Chloroplast Genomes in Chenopodium and Its Related Genera (Amaranthaceae): New Insights into Phylogenetic Relationships and the Development of Species-Specific Molecular Markers.

Species within the genus Chenopodium hold significant research interest due to their nutritional richness and salt tolerance. However, the morphological similarities among closely related species and a dearth of genomic resources have impeded their comprehensive study and utilization. In the present research, we conduct the sequencing and assembly of chloroplast (cp) genomes from six Chenopodium and related species, five of which were sequenced for the first time. These genomes ranged in length from 151,850 to 152,215 base pairs, showcased typical quadripartite structures, and encoded 85 protein-coding genes (PCGs), 1 pseudogene, 37 tRNA genes, and 8 rRNA genes. Compared with the previously published sequences of related species, these cp genomes are relatively conservative, but there are also some interspecific differences, such as inversion and IR region contraction. We discerned 929 simple sequence repeats (SSRs) and a series of highly variable regions across 16 related species, predominantly situated in the intergenic spacer (IGS) region and introns. The phylogenetic evaluations revealed that Chenopodium is more closely related to genera such as Atriplex, Beta, Dysphania, and Oxybase than to other members of the Amaranthaceae family. These lineages shared a common ancestor approximately 60.80 million years ago, after which they diverged into distinct genera. Based on InDels and SNPs between species, we designed 12 pairs of primers for species identification, and experiments confirmed that they could completely distinguish 10 related species.

Genome-wide identification of microsatellites for mapping, genetic diversity and cross-transferability in wheat (Triticum spp)

Wheat (Triticum aestivum L.) is a crucial global staple crop, and is consistently being improved to enhance yield, disease resistance, and quality traits. However, the development of molecular markers is a challenging task due to its hexaploid genome. Molecular marker system such as simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) are helpful for breeding, but SNP has limitations due to its development cost and its conversion to breeder markers. The study proposed an in-silico approach, by utilizing the low-cost transcriptome sequencing of two parental lines, ‘TAC 75′ and ‘WH 1105′, to identify polymorphic SSRs for mapping in a recombinant inbred line (RIL) population. This study introduces a new approach to bridge wheat genetics intricacies and next-generation sequencing potential. It presents a comprehensive genome-wide SSR distribution using IWGSC CS RefSeq v2.1 genome assembly and to identify 189 polymorphic loci through in-silico strategy. Of these, 54.76% showed polymorphism between parents, surpassing the traditional low polymorphic success rate. A RIL population screening validated these markers, demonstrating the fitness of identified markers through chi-square tests. The designed SSRs were also validated for genetic diversity analysis in a subset of 37 Indian wheat genotypes and cross-transferability in the wild/relative wheat species. In diversity analysis, a subset of 38 markers revealed 95 alleles (2.5 allele/locus), indicating substantial genetic variation. Population structure analysis unveiled three distinct groups, supported by phylogenetic and PCoA analyses. Further the polymorphic SSRs were also analyzed for SSR-gene association using gene ontology analysis. By utilizing the developing seed transcriptome data within parental lines, the study has enhanced the polymorphic SSR identification precision and facilitated in the RIL population. The undertaken study pioneers the use of transcriptome sequencing and genetic mapping to overcome challenges posed by the intricate wheat genome. This approach offers a cost-effective, less labour-intensive alternative to conventional methods, providing a platform for advancing wheat breeding research.

Genetic factors of early Alzheimer's dementia

Alzheimer's disease (AD) is classified as a neurodegenerative disorder that alters the memory and cognitive capacity of the affected individual, in addition to causing behavioral changes and other neuropsychiatric symptoms. In this work, genes involved in the metabolism of this neurodegenerative disorder were reported, as well as their positioning on their chromosomes with an emphasis on the human genome. The following research sources were used to collect data and results: scientific articles, books and websites specializing in this subject regarding AD and the human genome. The result of this investigation indicated that the genes studied are located on chromosomes 1, 14, 17, 19 and 21 in the human genome, and associated with early AD disease in humans. Therefore, the identification of genes related to AD is necessary for a better understanding of AD genetics and also for the development of molecular markers of therapeutic targets for the diagnosis and treatment of this dementia.

Chromosome-scale genomes of commercially important mahoganies, Swietenia macrophylla and Khaya senegalensis

Mahogany species (family Meliaceae) are highly valued for their aesthetic and durable wood. Despite their economic and ecological importance, genomic resources for mahogany species are limited, hindering genetic improvement and conservation efforts. Here we perform chromosome-scale genome assemblies of two commercially important mahogany species: Swietenia macrophylla and Khaya senegalensis. By combining 10X sequencing and Hi-C data, we assemble high-quality genomes of 274.49 Mb (S. macrophylla) and 406.50 Mb (K. senegalensis), with scaffold N50 lengths of 8.51 Mb and 7.85 Mb, respectively. A total of 99.38% and 98.05% of the assembled sequences are anchored to 28 pseudo-chromosomes in S. macrophylla and K. senegalensis, respectively. We predict 34,129 and 31,908 protein-coding genes in S. macrophylla and K. senegalensis, respectively, of which 97.44% and 98.49% are functionally annotated. The chromosome-scale genome assemblies of these mahogany species could serve as a vital genetic resource, especially in understanding the properties of non-model woody plants. These high-quality genomes could support the development of molecular markers for breeding programs, conservation efforts, and the sustainable management of these valuable forest resources.

Development of Molecular Markers to Detect Diaporthe spp. from Decayed Soybean Seeds

Soybean is one of the world’s most widely cultivated food crops, and soybean seeds are supplied from national seed resources in Korea. However, the transmission of seed-borne diseases through infected soybean seeds is problematic. Among these diseases, soybean seed decay is caused by Diaporthe spp. Infecting the pods, and the infected seeds show rotting symptoms. Most diseased seeds are removed during the selection process; however, it is difficult to distinguish infected seeds that do not display symptoms. Hence, a sequence-based method was devised to screen Diaporthe-infected seeds. Based on the nuclear ribosomal internal transcribe spacer (ITS) region of the pathogen, a primer was designed to distinguish the infection from other soybean seed pathogens. As a result of the comparison between healthy and Diaporthe-diseased seeds by using the primers, Diaporthe was detected only in the diseased seeds. Therefore, it is possible to distribute healthy soybean seeds by detecting Diaporthe-diseased seeds at the genetic level using the Diaporthe-specific primers.

De novo assembly of Iron-Heart Cunninghamia lanceolata transcriptome and EST-SSR marker development for genetic diversity analysis.

Iron-Heart Cunninghamia lanceolata, a wild relative of Chinese fir with valuable genetic and breeding traits, has been limited in genetic studies due to a lack of genomic resources and markers. In this study, we conducted transcriptome sequencing of Iron-Heart C. lanceolata leaves using Illumina NovaSeq 6000 and performed assembly and analysis. We obtained 45,326,576 clean reads and 115,501 unigenes. Comparative analysis in five functional databases resulted in successful annotation of 26,278 unigenes, with 6,693 unigenes annotated in all databases (5.79% of the total). UniProt and Pfam databases provided annotations for 22,673 and 18,315 unigenes, respectively. Gene Ontology analysis categorized 23,962 unigenes into three categories. KEGG database alignment annotated 10,195 unigenes, classifying them into five categories: metabolism, genetic information, biological systems, cellular processes, and environmental information processing. From the unigenes, we identified 5,645 SSRs, with dinucleotides repeats being the most common (41.47%). We observed variations in repeat numbers and base compositions, with the majority of markers ranging from 12 to 29 bp in length. We randomly selected 200 primer pairs and successfully amplified 15 pairs of polymorphic SSR primers, which effectively distinguished Chinese fir plants of different origins. This study provides insights into the genetic characteristics of Iron-Heart C. lanceolata and offers a foundation for future molecular marker development, breeding programs, genetic diversity analysis, and conservation strategies.

Development of EST-SSR molecular markers related to metabolic pathways in Kelussia odoratissima Mozaff

Development of EST-SSR molecular markers related to metabolic pathways in Kelussia odoratissima Mozaff

Genome-Wide Association Studies of Biluochun Tea Plant Populations in Dongting Mountain and Comprehensive Identification of Candidate Genes Associated with Core Agronomic Traits by Four Analysis Models.

The elite germplasm resources are key to the beautiful appearance and pleasant flavor of Biluochun tea. We collected and measured the agronomic traits of 95 tea plants to reveal the trait diversity and breeding value of Biluochun tea plant populations. The results revealed that the agronomic traits of Biluochun tea plant populations were diverse and had high breeding value. Additionally, we resequenced these tea plant populations to reveal genetic diversity, population structure, and selection pressure. The Biluochun tea plant populations contained two groups and were least affected by natural selection based on the results of population structure and selection pressure. More importantly, four non-synonymous single nucleotide polymorphisms (nsSNPs) and candidate genes associated with (-)-gallocatechin gallate (GCG), (-)-gallocatechin (GC), and caffeine (CAF) were detected using at least two GWAS models. The results will promote the development and application of molecular markers and the utilization of elite germplasm from Biluochun populations.

Comparative Analysis Revealed Intrageneric and Intraspecific Genomic Variation in Chloroplast Genomes of Actinidia spp. (Actinidiaceae, Viridiplantae)

About ten species of the genus Actinidia Lindl. are known as cultivated plants—alongside the most known A. chinensis var. deliciosa, there are many others, including Far-Eastern cold-hardy kiwifruits such as A. arguta, A. kolomikta, and A. polygama. Unlike most plant species, in which the chloroplast genome is maternally inherited, the family Actinidiaceae possesses a complex system of plastid inheritance with possible transmission through both maternal and paternal lines. The main aim of this work was the assembly of the plastid genomes of three species of Actinidia, their comparison with already-available sequences from databases, and evolutionary analyses. We discovered that the gene composition and gene sequences are conserved; the studied species are either subject to purifying selection or not subject to selection at all (with some exceptions, such as the ycf2 gene). However, the chloroplast chromosomes of some Actinidia species have undergone significant structural rearrangements, leading to the persistence of two main forms, both on an intrageneric and intraspecific level. These results expand our understanding of plastid genomics and genetic diversity within the genus Actinidia, providing a basis for future research in molecular marker development, phylogenetic analysis, and population studies.