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DArTseq-Based High-Throughput SilicoDArT and SNP Markers Applied for Association Mapping of Genes Related to Maize Morphology.

ABSTRACT: Today, agricultural productivity is essential to meet the needs of a growing population, and is also a key tool in coping with climate change. Innovative plant breeding technologies such as molecular markers, phenotyping, genotyping, the CRISPR/Cas method and next-generation sequencing can help agriculture meet the challenges of the 21st century more effectively. Therefore, the aim of the research was to identify single-nucleotide polymorphisms (SNPs) and SilicoDArT markers related to select morphological features determining the yield in maize. The plant material consisted of ninety-four inbred lines of maize of various origins. These lines were phenotyped under field conditions. A total of 14 morphological features was analyzed. The DArTseq method was chosen for genotyping because this technique reduces the complexity of the genome by restriction enzyme digestion. Subsequently, short fragment sequencing was used. The choice of a combination of restrictases allowed the isolation of highly informative low copy fragments of the genome. Thanks to this method, 90% of the obtained DArTseq markers are complementary to the unique sequences of the genome. All the observed features were normally distributed. Analysis of variance indicated that the main effect of lines was statistically significant (p < 0.001) for all 14 traits of study. Thanks to the DArTseq analysis with the use of next-generation sequencing (NGS) in the studied plant material, it was possible to identify 49,911 polymorphisms, of which 33,452 are SilicoDArT markers and the remaining 16,459 are SNP markers. Among those mentioned, two markers associated with four analyzed traits deserved special attention: SNP (4578734) and SilicoDArT (4778900). SNP marker 4578734 was associated with the following features: anthocyanin coloration of cob glumes, number of days from sowing to anthesis, number of days from sowing to silk emergence and anthocyanin coloration of internodes. SilicoDArT marker 4778900 was associated with the following features: number of days from sowing to anthesis, number of days from sowing to silk emergence, tassel: angle between the axis and lateral branches and plant height. Sequences with a length of 71 bp were used for physical mapping. The BLAST and EnsemblPlants databases were searched against the maize genome to identify the positions of both markers. Marker 4578734 was localized on chromosome 7, the closest gene was Zm00001d022467, approximately 55 Kb apart, encoding anthocyanidin 3-O-glucosyltransferase. Marker 4778900 was located on chromosome 7, at a distance of 45 Kb from the gene Zm00001d045261 encoding starch synthase I. The latter observation indicated that these flanking SilicoDArT and SNP markers were not in a state of linkage disequilibrium.

SUBMITTER: Tomkowiak A

PROVIDER: S-EPMC8198497 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Macadamia (Macadamia integrifolia, M. tetraphylla and hybrids) is an Australian native nut crop and has a significant economic value in the food industries worldwide. Long juvenility along with traditional breeding strategies impede quick genetic improvement of this crop. The existing cultivars constitute only second to fourth generation of the wild germplasm in the rainforest. The utilisation of molecular markers for genomic selection and genome-wide association studies may accelerate genetic gains. Identification of a robust, reproducible, and cost-effective marker system is instrumental in increasing the efficiency of genomic studies. This study is the first to report the potential of two ultra-high-throughput diversity array technology (DArT) markers (silicoDArT and SNP) in macadamia. Both markers were used to identify the genetic diversity and population structure in 80 macadamia cultivars. Parentage analysis of 25 scions in a rootstock trial was conducted to confirm plant identity where recorded identities did not corroborate with phenotypic field observations. A total of 22,280 silicoDArT and 7,332 SNP markers were reported, of which 11,526 silicoDArT and 3,956 SNP markers were used for analyses after screening with quality control parameters including >95% call rate, >95% reproducibility, and >0.05 one ratio. The average polymorphic information content (PIC) values of silicoDArT and SNP markers were 0.29 and 0.21, respectively. Genetic variance among the cultivars ranged from 0.003 to 0.738 in silicoDArT and 0.004 to 0.412 in SNP markers. Four distinct population groups were identified from SNP data analysis. Most of the accessions used in this study were descended from two or more populations. Cluster analysis clearly separated genotypes of distinct origins, such as the Hawaii Agricultural Experiment Station and Hidden Valley Plantation accessions. Two wild accessions of Macadamia jansenii and M. ternifolia were found to be distantly related to the cultivars. Wild germplasm individuals and their hybrids with cv. '660' formed separate clusters, suggesting that crossing between wild and cultivated genepools can extend genetic diversity. DArTseq-based SNP markers were successfully utilized to confirm the genetic identity of 25 scions in a rootstock trial. Our study suggests that DArT platforms are a robust system for the facilitation of genomic studies with regard to macadamia.

Project description:In the last decade, many scientists have used molecular biology methods in their research to locate the grain-yield-determining loci and yield structure characteristics in maize. Large-scale molecular analyses in maize do not only focus on the identification of new markers and quantitative trait locus (QTL) regions. DNA analysis in the selection of parental components for heterotic crosses is a very important tool for breeders. The aim of this research was to identify and select new markers for maize (SNP and SilicoDArT) linked to genes influencing the size of the yield components in maize. The plant material used for the research was 186 inbred maize lines. The field experiment was established in twolocations. The yield and six yield components were analyzed. For identification of SNP and SilicoDArT markers related to the yield and yield components, next-generation sequencing was used. As a result of the biometric measurements analysis, differentiation in the average elevation of the analyzed traits for the lines in both locations was found. The above-mentioned results indicate the existence of genotype-environment interactions. The analysis of variance for the observed quality between genotypes indicated a statistically significant differentiation between genotypes and a statistically significant differentiation for all the observed properties betweenlocations. A canonical variable analysis was applied to present a multi-trait assessment of the similarity of the tested maize genotypes in a lower number of dimensions with the lowest possible loss of information. No grouping of lines due to the analyzed was observed. As a result of next-generation sequencing, the molecular markers SilicoDArT (53,031) and SNP (28,571) were obtained. The genetic distance between the analyzed lines was estimated on the basis of these markers. Out of 81,602 identified SilicoDArT and SNP markers, 15,409 (1559 SilicoDArT and 13,850 SNPs) significantly related to the analyzed yield components were selected as a result of association mapping. The greatest numbers of molecular markers were associated with cob length (1203), cob diameter (1759), core length (1201) and core diameter (2326). From 15,409 markers significantly related to the analyzed traits of the yield components, 18 DArT markers were selected, which were significant for the same four traits (cob length, cob diameter, core length, core diameter) in both Kobierzyce and Smolice. These markers were used for physical mapping. As a result of the analyses, it was found that 6 out of 18 (1818; 14,506; 2317; 3233; 11,657; 12,812) identified markers are located inside genes. These markers are located on chromosomes 8, 9, 7, 3, 5, and 1, respectively.

Project description:The chief aim of plant breeding is to improve varieties so as to increase their yield and breeding traits. One of the first stages of breeding is the selection of parental forms from the available gene pool of existing varieties. To date, costly and laborious methods based on multiple crossbreeding and phenotypic selection have been necessary to properly assess genetic resources in terms of productivity, quality parameters, and susceptibility to biotic and abiotic stressors. The often long and complicated breeding cycle can be significantly shortened through selection using DNA markers. To this end, use is made of close couplings between the marker and the locus responsible for the inheritance of the functional trait. The aim of this study was to identify single nucleotide polymorphism (SNP) and SilicoDArT markers associated with yield traits and to predict the heterosis effect for yield traits in maize (Zea mays L.). The plant material used in the research consisted of 19 inbred maize lines derived from different starting materials, and 13 hybrids resulting from crossing them. A two-year field experiment with inbred lines and hybrids was established at two Polish breeding stations on 10 m2 plots in a randomized block design with three replicates. The biometric measurements included cob length, cob diameter, core length, core diameter, number of rows of grain, number of grains in a row, mass of grain from the cob, weight of one thousand grains, and yield. The isolated DNA was subjected to DArTseq genotyping. Association mapping was performed in this study using a method based on the mixed linear model with the population structure estimated by eigenanalysis (principal component analysis of all markers) and modeled by random effects. Narew, Popis, Kozak, M Glejt, and Grom were the hybrids used in the study that showed the highest significant heterosis effect in 2013 and 2014. The similarity between parental components determined on the basis of SNP and SilicoDArT marker analysis did not exceed 33%. It was found that the genetic similarity between parental components, determined on the basis of SNP and SilicoDArT markers, reflected their degree of relationship, and correlated significantly with the effect of heterosis. As the results indicate, the parental components for heterosis crosses can be selected based on genetic similarity between parental components evaluated using SNP and SilicoDArT markers, supported with information on the origin of parental forms. Of the markers we analyzed, 76 were selected as being significantly associated with at least six traits observed in 2013 and 2014 at both the Łagiewniki and Smolice stations.

Project description:Establishing the genetic diversity and population structure of a species can guide the selection of appropriate conservation and sustainable utilization strategies. Next-generation sequencing (NGS) approaches are increasingly being used to generate multi-locus data for genetic structure determination. This study presents the genetic structure of a fodder species -Trema orientalis based on two genome-wide high-throughput diversity array technology (DArT) markers; silicoDArT and single nucleotide polymorphisms (SNPs). Genotyping of 119 individuals generated 40,650 silicoDArT and 4767 SNP markers. Both marker types had a high average scoring reproducibility (>99%). Genetic relationships explored by principal coordinates analysis (PCoA) showed that the first principal coordinate axis explained most of the variation in both the SilicoDArT (34.2%) and SNP (89.6%) marker data. The average polymorphic information content did not highly differ between silicoDArT (0.22) and SNPs (0.17) suggesting minimal differences in informativeness in the two groups of markers. The, mean observed (Ho) and expected (He) heterozygosity were low and differed between the silicoDArT and SNPs respectively, estimated at Ho = 0.08 and He = 0.05 for silicoDArT and Ho = 0.23 and He = 0.19 for SNPs. The population of T. orientalis was moderately differentiated (FST = 0.20-0.53) and formed 2 distinct clusters based on maximum likelihood and principal coordinates analysis. Analysis of molecular variance revealed that clusters contributed more to the variation (46.3-60.8%) than individuals (32.9-31.2%). Overall, the results suggest a high relatedness of the individuals sampled and a threatened genetic potential of T. orientalis in the wild. Therefore, genetic management activities such as ex-situ germplasm management are required for the sustainability of the species. Ex-situ conservation efforts should involve core collection of individuals from different populations to capture efficient diversity. This study demonstrates the importance of silicoDArT and SNP makers in population structure and genetic diversity analysis of Trema orientalis, useful for future genome wide studies in the species.

Project description:Cassava (Manihot esculenta Crantz) is an important industrial and staple crop due to its high starch content, low input requirement, and resilience which makes it an ideal crop for sustainable agricultural systems and marginal lands in the tropics. However, the lack of genomic information on local genetic resources has impeded efficient conservation and improvement of the crop and the exploration of its full agronomic and breeding potential. This work was carried out to obtain information on population structure and extent of genetic variability among some local landraces conserved at the Plant Genetic Resources Research Institute, Ghana and exotic cassava accessions with Diversity Array Technology based SilicoDArT and SNP markers to infer how the relatedness in the genetic materials can be used to enhance germplasm curation and future breeding efforts. A total of 10521 SilicoDArT and 10808 SNP markers were used with varying polymorphic information content (PIC) values. The average PIC was 0.36 and 0.28 for the SilicoDArT and SNPs respectively. Population structure and average linkage hierarchical clustering based on SNPs revealed two distinct subpopulations and a large number of admixtures. Both DArT platforms identified 22 landraces as potential duplicates based on Gower's genetic dissimilarity. The expected heterozygosity which defines the genetic variation within each subpopulation was 0.008 for subpop1 which were mainly landraces and 0.391 for subpop2 indicating the homogeneous and admixture nature of the two subpopulations. Further analysis upon removal of the duplicates increased the expected heterozygosity of subpop1 from 0.008 to 0.357. A mantel test indicated strong interdependence (r = 0.970; P < 0.001) between SilicoDArT and DArTSeq SNP genotypic data suggesting both marker platforms as a robust system for genomic studies in cassava. These findings provide important information for efficient ex-situ conservation of cassava, future heterosis breeding, and marker-assisted selection (MAS) to enhance cassava improvement.

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DArTseq-Based High-Throughput SilicoDArT and SNP Markers Applied for Association Mapping of Genes Related to Maize Morphology.

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