Biological responses of an elite centipedegrass [Eremochloa ophiuroides (Munro) Hack.] cultivar (Ganbei) to carbon ion beam irradiation.

Carbon ion beam irradiation (CIBI) is a highly efficient mutagenesis for generating mutations that can be used to expand germplasm resources and create superior new germplasm. The study investigated the effects of different doses of CIBI (50 Gy, 100 Gy, 150 Gy, 200 Gy and 300 Gy) on seed germination and seedling survival, seedling morphological and physiological traits of an elite centipedegrass cultivar Ganbei. The results showed that irradiation greater than 50 Gy cause inhibition of seed germination, and the semi-lethal dose (LD50) is around 90 Gy for CIBI treated seeds of Ganbei. A carbon ion beam-mutagenized centipedegrass population was generated from Ganbei, with irradiation dosages from 50 Gy to 200 Gy. More than ten types of phenotypic variations and novel mutants with heritable tendencies mainly including putative mutants of stolon number, length and diameter, of internode length, of leaf length and width, of leaf chlorophyll content, of stolon growth rate, of aboveground tissue dry weight, of sward height were identified. While the total sugar content of the plants from irradiated seeds showed no obvious change in all treatments as compared to the control, the crude protein content displayed significant reduction at a high-dose treatment of 200 Gy. Genetic polymorphism was detected in mutagenized centipedegrass population using SSR-PCR analysis, suggesting that CIBI caused alteration of larger fragments of the DNA sequence. As a result, a preliminary batch of mutants was screened in this study. In summary, carbon ion beam mutagenesis is an effective way for developing centipedegrass germplasm with wider variation, and treating seeds with CIBI at a dosage of ~100 Gy could be effective in centipedegrass mutation breeding.

Morphological, biochemical and molecular characterization of short-day tropical Indian garlic (Allium sativum L.).

Garlic, an asexually propagated bulbous crop, displays a wide diversity based on its morphological traits and biochemical compositions. This study investigated the genetic variability of Indian garlic through morphological, biochemical, and molecular markers. Twenty-nine genotypes along with three Allium species as outgroup were included in the present study. Observations were recorded on 14 quantitative traits, 17 qualitative traits, and 9 biochemical traits in fresh garlic. Significant variability was observed among genotypes for different characters. All the morphological and biochemical traits showed higher phenotypic coefficient of variation (PCV) than genotypic coefficient of variation (GCV) revealing the role of environment in trait expression. High to moderate heritability and genetic advance as percent mean were recorded for different traits except dry matter and Total Soluble Solids (TSS). Correlation analysis revealed the highest positive correlation between total yield, marketable yield, Ferric Reducing Antioxidant Potential (FRAP) and 2,2-diphenyl-1-picrylhyrazyl (DPPH). Cluster analysis differentiated all the genotypes into three major clusters based on morphological and biochemical traits. 214 Simple Sequence Repeats (SSRs) were screened and nine markers exhibited polymorphism. Cluster analysis using molecular markers revealed 4 distinct clusters. The observations from this study will help in the identification of diverse garlic germplasm for its efficient management and duplicate identification of germplasm resources.

Assessment of Population Genetic Diversity of Medicinal Meconopsis integrifolia (Maxim.) Franch. Using Newly Developed SSR Markers.

Meconopsis integrifolia is an endangered Tibetan medicinal plant with significant medicinal and ornamental value. Understanding its genetic diversity and structure is crucial for its sustainable utilization and effective conservation. Here, we develop a set of SSR markers based on transcriptome data to analyze the genetic diversity and structure of 185 individuals from 16 populations of M. integrifolia. The results indicate that M. integrifolia exhibits relatively high genetic diversity at the species level (the percentage of polymorphic bands PPB = 91.67%, Nei's genetic diversity index He = 0.2989, Shannon's information index I = 0.4514) but limited genetic variation within populations (PPB = 12.08%, He = 0.0399, I = 0.0610). The genetic differentiation among populations is relatively high (the coefficient of gene differentiation GST = 0.6902), and AMOVA analysis indicates that 63.39% of the total variation occurs among populations. This suggests that maintaining a limited number of populations is insufficient to preserve the overall diversity of M. integrifolia. Different populations are categorized into four representative subclusters, but they do not cluster strictly according to geographical distribution. Limited gene flow (Nm = 0.2244) is likely the main reason for the high differentiation among these populations. Limited seed and pollen dispersal abilities, along with habitat fragmentation, may explain the restricted gene flow among populations, highlighting the necessity of conserving as many populations in the wild as possible.

Streamlining of Simple Sequence Repeat Data Mining Methodologies and Pipelines for Crop Scanning.

Genetic markers are powerful tools for understanding genetic diversity and the molecular basis of traits, ushering in a new era of molecular breeding in crops. Over the past 50 years, DNA markers have rapidly changed, moving from hybridization-based and second-generation-based to sequence-based markers. Simple sequence repeats (SSRs) are the ideal markers in plant breeding, and they have numerous desirable properties, including their repeatability, codominance, multi-allelic nature, and locus specificity. They can be generated from any species, which requires prior sequence knowledge. SSRs may serve as evolutionary tuning knobs, allowing for rapid identification and adaptation to new circumstances. The evaluations published thus far have mostly ignored SSR polymorphism and gene evolution due to a lack of data regarding the precise placements of SSRs on chromosomes. However, NGS technologies have made it possible to produce high-throughput SSRs for any species using massive volumes of genomic sequence data that can be generated fast and at a minimal cost. Though SNP markers are gradually replacing the erstwhile DNA marker systems, SSRs remain the markers of choice in orphan crops due to the lack of genomic resources at the reference level and their adaptability to resource-limited labor. Several bioinformatic approaches and tools have evolved to handle genomic sequences to identify SSRs and generate primers for genotyping applications in plant breeding projects. This paper includes the currently available methodologies for producing SSR markers, genomic resource databases, and computational tools/pipelines for SSR data mining and primer generation. This review aims to provide a 'one-stop shop' of information to help each new user carefully select tools for identifying and utilizing SSRs in genetic research and breeding programs.

Genetic diversity evaluation of Luculia yunnanensis, a vulnerable species endemic to Yunnan, Southwestern China based on morphological traits and EST-SSR markers.

Luculia yunnanensis is a vulnerable species endemic to Yunnan Province, Southwestern China, which has high ornamental value. Its wild population has not been fully protected and utilized for a long time, which is not conducive to the long-term stable development of this species. Genetic diversity assessment is the basis and prerequisite for the conservation of rare species. In this study, 21 phenotypic traits and 17 highly polymorphic EST-SSR markers were used to analyze the genetic diversity and genetic structure of 164 individuals from six L. yunnanensis populations. The coefficient of variation of 21 phenotypic traits ranged from 11.76% to 52.58% (mean=21.72%), and the coefficient of variation of 18 traits was less than 30%. The average values of Ne, I, Ho and He were 1.710, 0.619, 0.384, and 0.352, respectively. The genetic diversity of LLO (Ho = 0.476 and He = 0.426) and LCM (Ho = 0.424 and He = 0.381) populations in Lushui County was highest. The GDX populations (Ho = 0.335 and He = 0.269) isolated by Gaoligong Mountain had the lowest genetic diversity. The AMOVA results showed that 13.04% of the genetic variation was among populations and 86.96% was within populations. The average phenotypic differentiation coefficient of phenotypic traits among populations was 18.69%. The results of phenotypic and genetic variation analysis were consistent, indicating that the most of variation exists within population. Genetic structure, UPGMA clustering and PCA analysis results showed that the populations of L. yunnanensis had obvious geographical divisions, and the populations distributed in the southern region and distributed in the northern region of the Nujiang River clustered into one group respectively. Combining the results of phenotypic and molecular markers, we recommend that give priority to the protection of LLO, LCM and GDX population, in order to ensure the sustainable utilization of L. yunnanensis germplasm resources.

Association mapping analysis (AMA) for morpho-agronomic traits and leaf aromatic compounds using SSR markers in three types of Perilla crop collected from South Korea.

BACKGROUND: Perilla is a representative leafy vegetable in South Korea. As K-Food (Korean food) is in the spotlight around the world, there is also increasing interest in Western countries in Perilla crop, an annual plant belonging to the Lamiaceae family. OBJECTIVE: To discover comprehensive information, including genetic and phylogenetic relationships among the 80 native Perilla accessions, using three types of data: simple sequence repeat (SSR) marker data, volatiles profile data, and morpho-agronomic data. METHODS: This study conducted genotypic and phenotypic analyses on 80 Perilla accessions of three types (cultivated var. frutescens, weedy var. frutescens, weedy var. crispa) from South Korea. Five groups (G1-G5) of the 80 Perilla accessions of the three types were differentiated into two different clusters [genotype-based clustering (GTC) and phenotype-based clustering (PTC)] based on an aroma sensory phenotypic test. RESULTS: A total of 314 alleles were confirmed using 55 Perilla SSR primer sets, and genetic variation in the 80 Perilla accessions was evaluated. Among the three statistical analysis methods, principal coordinate analysis (PCoA) and GTC using data of the 55 Perilla SSR markers revealed perfectly consistent results, whereas PTC produced a total of six clusters. The 10 Perilla SSR markers associated with and significantly correlated with both biochemical and morphological characteristics were selected. CONCLUSIONS: These findings are expected to provide valuable information for developing global South Korean Perilla cultivars for further studies in Perilla crop breeding programs.

Genetic markers identify duplicates in Nordic potato collections.

Introduction: The first small scale cultivation of potatoes in the Nordic countries began roughly 300 years ago, and later became an important staple food in the region. Organized conservation efforts began in the 1980s, and today, potato landraces, improved varieties, and breeding lines are conserved in genebanks at the Nordic Genetic Resource Center (NordGen), Sweden, and the Norwegian Genetic Resource Centre (NGS), Norway, as well as at potato breeding companies across Nordic countries. All these collections house a diverse array of genotypes with local names and local growing histories from the whole region. However, the presence of duplicates, and inconsistent naming has led to confusion. Methods: In this study, 198 accessions of cultivated potato (Solanum tuberosum L.) have been genotyped with 62 microsatellite (SSR) markers. The analyzed accessions came from three collections: 43 accessions from the Danish Potato Breeding Foundation in Vandel (LKF-Vandel), 90 from NordGen and 65 from NGS. Results and discussion: The genetic analysis revealed 140 unique potato genotypes and 31 groups/clusters of duplicates, most of which contained duplicate pairs and the others three to ten accessions. Several accessions with distinct names were genetically identical or very similar, suggesting historical sharing, and regional distribution of seed potatoes, leading to the emergence of diverse local names. Moreover, many improved varieties from early potato breeding were revealed to have duplicates that have been considered Nordic landraces. Furthermore, potato accessions with identical names but originating from different collections were confirmed to be duplicates. These findings have already influenced management decisions and will further improve management practices for Nordic potato collections. Additionally, this new knowledge will benefit Nordic potato breeding efforts and allow for the dissemination of more accurate information to other users of potato diversity.

Genetic diversity analysis and DNA fingerprint construction of Zanthoxylum species based on SSR and iPBS markers.

Zanthoxylum is a versatile economic tree species utilized for its spice, seasoning, oil, medicinal, and industrial raw material applications, and it has a lengthy history of cultivation and domestication in China. This has led to the development of numerous cultivars. However, the phenomenon of mixed cultivars and confusing names has significantly obstructed the effective utilization of Zanthoxylum resources and industrial development. Consequently, conducting genetic diversity studies and cultivar identification on Zanthoxylum are crucial. This research analyzed the genetic traits of 80 Zanthoxylum cultivars using simple sequence repeat (SSR) and inter-Primer Binding Site (iPBS) molecular markers, leading to the creation of a DNA fingerprint. This study identified 206 and 127 alleles with 32 SSR markers and 10 iPBS markers, respectively, yielding an average of 6.4 and 12.7 alleles (Na) per marker. The average polymorphism information content (PIC) for the SSR and iPBS markers was 0.710 and 0.281, respectively. The genetic similarity coefficients for the 80 Zanthoxylum accessions ranged from 0.0947 to 0.9868 and from 0.2206 to 1.0000, with mean values of 0.3864 and 0.5215, respectively, indicating substantial genetic diversity. Cluster analysis, corroborated by principal coordinate analysis (PCoA), categorized these accessions into three primary groups. Analysis of the genetic differentiation among the three Zanthoxylum (Z. bungeanum, Z. armatum, and Z. piperitum) populations using SSR markers revealed a mean genetic differentiation coefficient (Fst) of 0.335 and a gene flow (Nm) of 0.629, suggesting significant genetic divergence among the populations. Molecular variance analysis (AMOVA) indicated that 65% of the genetic variation occurred within individuals, while 35% occurred among populations. Bayesian model-based analysis of population genetic structure divided all materials into two groups. The combined PI and PIsibs value of the 32 SSR markers were 4.265 × 10- 27 and 1.282 × 10- 11, respectively, showing strong fingerprinting power. DNA fingerprints of the 80 cultivars were established using eight pairs of SSR primers, each assigned a unique numerical code. In summary, while both markers were effective at assessing the genetic diversity and relationships of Zanthoxylum species, SSR markers demonstrated superior polymorphism and cultivar discrimination compared to iPBS markers. These findings offer a scientific foundation for the conservation and sustainable use of Zanthoxylum species.

In silico EST-SSR Identification and Development through EST Sequences from Metroxylon sagu Rottb. for Genetic Diversity Analysis.

Sago plant (Metroxylon sagu Rottb.) is one of the most carbohydrate-producing plants in the world. Microsatellites or simple sequence repeats (SSRs) play an important role in the genome and are used extensively compared to other molecular markers. For the first time, we are exploiting data expressed sequence tags (EST) of sago plants to identify and characterise markers in this species. EST data about sago plants are obtained through the EST database on the National Center for Biotechnology Information (NCBI) website. We obtained data of 458 Kb (412 contig) with a maximum and minimum length of 1,138 and 124 nucleotides, respectively. We successfully identified 820 perfectly patterned SSR using Phobos 3.3.12 software. The type characterisation of EST-SSR was dominated by tri-nucleotides 36% (294), followed by hexa-nucleotides 24% (202), tetra-nucleotides 15% (120), penta-nucleotides 13% (108) and di-nucleotides 12% (96). The most frequency of SSR motifs in each type is AG, AAG and AAAG. Analysis of synteny on the EST sequence with the online application Phytozome found that sequences were distributed on 12 Oryza sativa chromosomes with a likeness percentage between 63% to 100% and e-value between 0 to 0.094. We developed the primer and generated 19 primers. Furthermore, we validated 7 primers that all generated polymorphic alleles. To our knowledge, this report is the first identification and characterisation of EST-SSR for sago species and these markers can be used for genetic diversity analysis, marker assisted selection (MAS), cultivar identification, kinship analysis and genetic mapping analysis.

The Migration, Diversity, and Evolution of Puccinia triticina in China.

Wheat leaf rust, caused by Puccinia triticina, is one of the most common fungal diseases of wheat in China and occurs widely in various wheat-growing regions. To clarify the epidemic, spread rules, and population structure of P. triticina among different regions, 217 isolates of P. triticina collected from Hebei, Shandong, Sichuan, and Xinjiang in China were tested by 34 Thatcher near-isogenic lines and 21 pairs of EST-SSR primers. A total of 83 races were identified, and THTT, PHTT, THTS, and PHJT were the most predominant races in the four provinces in 2009. We found enriched virulence and genetic diversity in the four P. triticina populations and a significant correlation between genetic polymorphism and geographic regions. However, no significant correlation was found between virulence phenotypes and molecular genotypes. Moreover, a notable high level of gene flow (Nm = 2.82 > 1) among four P. triticina populations was detected. The genetic relationship among Hebei, Shandong, and Sichuan populations was close, possibly due to the spread of P. triticina from Sichuan to Shandong and then to Hebei. In contrast, the Xinjiang population was relatively independent. Genetic differentiation analysis showed some level of differentiation among or within populations of P. triticina in the four provinces, and the genetic variation within populations (74.97%) was higher than across populations (25.03%). Our study provides a basis for a better understanding of the regional migration, epidemic, and population structure of P. triticina in China.