Microsatellite Research Articles

Genetic Diversity of Tulipa alberti and T. greigii Populations from Kazakhstan Based on Application of Expressed Sequence Tag Simple Sequence Repeat Markers.

The genus Tulipa L., renowned for its ornamental and ecological significance, encompasses a diversity of species primarily concentrated in the Tian Shan and Pamir-Alay Mountain ranges. With its varied landscapes, Kazakhstan harbors 42 Tulipa species, including the endangered Tulipa alberti Regel and Tulipa greigii Regel, which are critical for biodiversity yet face significant threats from human activities. This study aimed to assess these two species' genetic diversity and population structure using 15 expressed sequence tag simple sequence repeat (EST-SSR) markers. Leaf samples from 423 individuals across 23 natural populations, including 11 populations of T. alberti and 12 populations of T. greigii, were collected and genetically characterized using EST-SSR markers. The results revealed relatively high levels of genetic variation in T. greigii compared to T. alberti. The average number of alleles per locus was 1.9 for T. alberti and 2.8 for T. greigii. AMOVA indicated substantial genetic variation within populations (75% for T. alberti and 77% for T. greigii). The Bayesian analysis of the population structure of the two species indicated an optimal value of K = 3 for both species, splitting all sampled populations into three distinct genetic clusters. Populations with the highest level of genetic diversity were identified in both species. The results underscore the importance of conserving the genetic diversity of Tulipa populations, which can help develop strategies for their preservation in stressed ecological conditions.

Micropropagation, micromorphological evaluation and genetic homogeneity validation of Cucumis collosus (Rottl.) Cogn.: A wild progenitor of melon

Cucumis collosus (Rottl.) Cogn., a wild ancestor of melon cultivars, is an underutilized cucurbit of arid horticulture having nutraceutical and therapeutic significance. C. collosus has great potential for enhancing the genetic base of cultivated melon and other cucurbits. This investigation is the first report on a robust and reliable micropropagation system for C. collosus (var. AHK-119) using nodal explants, followed by micromorphological and genetic stability assessment. More than 95% explants showed axillary shoot-bud induction on MS (Murashige and Skoog, 1962) medium containing 0.5 mg L−1 BAP and developed a maximum of 3.2 ± 0.6 shoots measuring 3.2 ± 0.1 cm length. After evaluating the various parameters, the best shoot multiplication (Shoot Number 34.9 ± 1.7, Shoot Length 6.7 ± 0.9 cm) was achieved on MS medium having half-strength of nitrates (825 mg L−1 NH4NO3 and 950 mg L−1 KNO3) + 0.25 mg L−1 6-benzylaminopurine (BAP) + 0.1 mg L−1 Kinetin (Kin) + 0.1 mg L−1 Indole-3-acetic acid (IAA) + additives (50 mg L−1 ascorbic acid and 25 mg L−1 each of adenine sulfate, L-arginine and citric acid), after 4 weeks. Micro-shoots were rooted using three approaches: In Vitro Continuous Treatment (IVCT), In Vitro Pulse-Treatment (IVPT) and Ex Vitro Pulse Treatment (EVPT). Of these, EVPT approach proved to be most successful in terms of rooting response (80%), root number (6.4 ± 0.9) and root length (5.7 ± 0.7 cm). The foliar micro-morphology analysis of in vitro leaves (IVL), ex vitro leaves (EVL) and mother plant leaves (MPL) demonstrated notable adaptive alterations during the transitional stages and produced structurally stable plants similar to the mother plant. Furthermore, the validation of genetic homogeneity using Start Codon Targeted (SCoT) and Inter Simple Sequence Repeats (ISSR) markers revealed monomorphic banding patterns among the micropropagated and the mother plant. The discussed method can be applied for large-scale commercial production of C. collosus for arid horticulture, urban/vertical farming, breeding purposes and improvement of other cucurbits.

Morphological and genetic characterization of mutants of Gladiolus cultivar prince of orange for two successive vegetative generations

Gladiolus mutants developed using gamma irradiations and EMS treatments were characterized using morphological, genetic and Polymerase Chain Reactions (PCR) based Inter-Simple Sequence Repeats (ISSR) for designing future breeding strategies. As many as 7 mutants based on phenotypic variations such as colour and shape modifications were subjected to molecular characterization using 11 ISSR primers that confirmed variations. ISSR markers showed 13.88 to 36.00 percent polymorphism among the mutants was recorded, signifying variability among different mutants. UPGMA (unweighted pair group method with arithmetic mean) based analysis of cluster separated gladiolus variety prince of orange mutants into four clusters. ISSR markers were subjected for comparative analysis like effective multiplex ratio, marker index, polymorphic information content and resolving power wherein, highest EMR (3.24), MI (1.32) were found for UBC 810, Rp (4.37) values were found for UBC 827, and maximum PIC value (0.46) for ISSR 22218 UBC 812. The results clearly indicated the reproducibility of ISSRs and their ability to detect variability among the mutants. A significant level of genetic variation was depicted by ANOVA which suggested the usefulness of studies in gladiolus breeding and unique bands obtained could be utilized for mutant/ varietal identification.

Relationship Between Heterosis and Genetic Distance Determined by SSR Markers in Oriental Tobacco

Hybrid cultivars could be beneficial to achieve high leaf yields while maintaining good quality properties in oriental tobacco. Identification of parents constitutes a major part of a hybrid breeding program. The aim of the present study was to determine the relationship between the genetic distance of parents determined by simple sequence repeats (SSR) markers and heterosis levels in hybrids produced from those parents for leaf yield and quality properties in oriental tobacco. Twenty-one hybrids produced by half diallel crossing among seven tobacco genotypes used in oriental tobacco production in Turkey were grown along with their parents in three locations in 2012 and 2013. Twenty-nine SSR markers were used to determine genetic distances among seven tobacco genotypes. A total of 80 alleles were produced by all twenty-nine markers. Average number of observed alleles per polymorphic marker was 2.96. Twenty-seven of 80 alleles were observed in only one of the seven parents. The polymorphic information content of markers varied from 0.215 to 0.810 (average 0.480). Mid-parent heterosis levels ranged from 18.03 to 42.00% for leaf number, between -19.75 and 38.06% for leaf width, between -17.51 and 36.25% for leaf length, between -34.38 and 76.12% for leaf yield, between -78.30 and 154.01% for sugar content and between -45.40 and 143.29% for nicotine content. Heterosis levels were correlated with genetic distances between parents for leaf number in Erbaa 2012 and Tokat 2012 locations, for leaf width in Erbaa 2013 location, for leaf length in Erbaa 2013 location, and for leaf yield in Tokat 2012 location only. SSR markers were very effective to determine genetic distance of oriental tobacco, and only two markers could distinguish all seven genotypes used in the study. The findings indicated that genetic distance determined by SSR markers used in the present study is not sufficient to predict hybrid performance in oriental tobacco.

Rpv10.2: A Haplotype Variant of Locus Rpv10 Enables New Combinations for Pyramiding Downy Mildew Resistance Traits in Grapevine.

In viticulture, pathogens like the oomycete Plasmopara viticola, the causal agent of downy mildew, can cause severe yield loss and require extensive application of plant protection chemicals. Breeders are generating pathogen-resistant varieties exploiting American and Asian wild Vitis germplasm as sources of resistance. Several loci mediating resistance to P. viticola have been identified in the past but may be overcome by specifically adapted strains of the pathogen. Aiming to find and characterize novel loci, a cross population with Vitis amurensis ancestry was investigated searching for resistance-correlated quantitative trait loci (QTL). As a prerequisite, a genetic map was generated by analyzing the 244 F1 individuals derived from a cross of the downy mildew susceptible Vitis vinifera cultivar 'Tigvoasa' and the resistant V. amurensis pBC1 breeding line We 90-06-12. This genetic map is based on the information from 627 molecular markers including 56 simple sequence repeats and 571 rhAmpSeq markers. A phenotypic characterization of the progeny showed a clear segregation of the resistance traits in the F1 population after an experimental inoculation of leaf discs with downy mildew. Combining genetic and phenotypic data, an analysis for QTL revealed a major locus on linkage Group 9 that correlates strongly with the resistance to downy mildew. The locus was mapped to a region of about 80 kb on the PN40024 (12x.V2) grapevine reference genome. This genomic region co-localizes with the formerly identified locus Rpv10 from the grapevine cultivar 'Solaris'. As we found different allele sizes of the locus-linked SSR markers than those characterizing the known Rpv10 locus and differences in the sequence of a candidate gene, it was regarded as a haplotype variant and named Rpv10.2.

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.

Production of genetically stable and Odontoglossum ringspot virus-free Cymbidium orchid ‘New True’ plants via meristem-derived protocorm-like body (PLB) subcultures

BackgroundThis study aimed to produce Odontoglossum ringspot virus (ORSV)-free Cymbidium orchid ‘New True’ plants from ORSV-infected mother plants by culturing their meristems and successively repeating subcultures of protocorm-like bodies (PLBs) derived from the meristems.ResultsInitially, ORSV was confirmed as the causative agent of viral symptoms in orchid leaves via reverse transcription-polymerase chain reaction (RT-PCR) analysis. Meristems from infected plants were cultured to generate PLBs, which in sequence were repeatedly subcultured up to four times. RT-PCR and quantitative RT-PCR analyses revealed that while ORSV was undetectable in shoots derived from the first subculture, complete elimination of the virus required at least a second subculture. Genetic analysis using inter-simple sequence repeat markers indicated no somaclonal variation between regenerated plants and the mother plant, suggesting that genetic consistency was maintained.ConclusionOverall, our findings demonstrate that subculturing PLBs for a second time is ideal for producing genetically stable, ORSV-free Cymbidium orchids, thus offering a practical means of generating genetically stable, virus-free plants and enhancing plant health and quality in the orchid industry.

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.

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. 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. 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. 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. These findings are expected to provide valuable information for developing global South Korean Perilla cultivars for further studies in Perilla crop breeding programs.

Population data for 12 X-STRs loci in Malaysian Malay and Chinese populations

The utilization of X-chromosome short tandem repeats (X-STRs) for human identification particularly in resolving complex kinship cases has been advocated. Since, forensic statistical parameters vary among different populations, and because the X-STRs population data representing the diverse population of Peninsular Malaysia remain unavailable, the specific attempt reported here for the Malays (n = 224) and Chinese (n = 201) populations appears forensically relevant to support the evidential value of the 12 X-STRs markers for human identification in Malaysia. Results derived from the Qiagen Investigator® Argus X-12 kit revealed that DXS10135 as the most polymorphic locus with high genetic diversity, polymorphic information content, heterozygosity as well as power of exclusion. Based on allele frequencies, the combined power of discrimination as well as the mean exclusion chance (MECKrüger, MECKishida, MECDesmarais and MECDesmaraisDuo) values for the Malays and Chinese were individually ≥0.999995532964908. As for the combined power of discrimination as well as the mean exclusion chance (MECKrüger, MECKishida, MECDesmarais and MECDesmaraisDuo) calculated based on haplotype frequencies, the values were ≥0.9999986410567 for the Malays and Chinese populations. In addition, results from the genetic distance, neighbor-joining phylogenetic tree and principal component analysis revealed close biogeographical distributions of the studied populations with other South East Asian populations. Hence, the utilization of the X-STRs data for identifying individuals among the Malays and Chinese populations in Peninsular Malaysia for forensic application appears as highly supported.

Efficient Plantlet Regeneration from Branches in Mangifera indica L.

Mango (Mangifera indica L.) is one of the most significant tropical and subtropical fruit species, with high ecological and economic value. However, research on the in vitro culture of mangoes is relatively weak, so establishing an efficient and stable mango plant regeneration system is of great significance. In this study, a preliminary mango regeneration system was established with Mangifera indica L. cv. Keitt from young branches as the starting explants. The results showed that the optimal plant growth regulator (PGR) formula for direct adventitious shoot induction on the branches was 1 mg/L 6-benzylaminopurine (6-BA) + 0.1 mg/L a-naphthaleneacetic acid (NAA), with an adventitious shoot induction rate of 73.63% and an average of 6.76 adventitious shoots. The optimal basal medium for adventitious shoot induction was wood plant medium (WPM), with an adventitious shoot induction rate of 63.87% and an average of 5.21 adventitious shoots. The optimal culture medium for adventitious shoot elongation was WPM + 1 mg/L 6-BA + 0.5 mg/L NAA, with an adventitious shoot elongation rate of 89.33% and an average length of 5.17 cm. The optimal formula for the induction of mango rooting was Douglas fir cotyledon revised medium (DCR) + 3 mg/L indole-3-butyric acid (IBA), with a maximum rooting rate of 66.13% and an average rooting quantity of 6.43. The genetic fidelity of the in vitro-regenerated plants was evaluated using inter-simple sequence repeat (ISSR) molecular markers. There was no difference between the in vitro-regenerated plants and the parent plant. This study provides an efficient and stable propagation system for Mangifera indica L., laying the foundation for its rapid propagation and genetic improvement.

Complete chloroplast genome of a montane plant Spathoglottis aurea Lindl.: Comparative analyses and phylogenetic relationships among members of tribe collabieae.

The yellow-flowered Spathoglottis aurea (tribe Collabieae; family Orchidaceae) is native to the mountainous areas of Peninsular Malaysia. The species is well known as an ornamental plant and for its role in artificial hybrid breeding. There is an interesting evolutionary relationship between S. aurea and the geographically isolated S. microchilina from Borneo that has encouraged further study of the S. aurea populations, but the genomic resource for S. aurea has not yet been reported. The present study reports the first work to characterize a chloroplast (cp) genome among the Spathoglottis genus. The complete cp genome of S. aurea was assembled from a sequence generated by the Illumina platform and analysed in comparison with other Collabieae species available in the GenBank database. The cp genome of S. aurea is 157,957 base pairs (bp) in length with guanine-cytosine (GC) content of 37.3%. The genome possessed a typical quadripartite cp genome structure with large single-copy (LSC) (86,888 bp), small single-copy (SSC) (18,125 bp) and inverted repeat (IR) (26,472 bp) sequences. A total of 134 genes were annotated, with 88 protein coding genes (PCGs), 38 transfer RNA (tRNA) genes and eight ribosomal RNA (rRNA) genes. Overall, 80 simple sequence repeats (SSR) or microsatellites were identified. Comparative analysis with other Collabieae species revealed high conservation in the cp genome arrangements with minimal difference in genome lengths. However, several mutational hotspots were also detected, with high potential to be developed as genetic markers for phylogenetic analysis. Characterization of the S. aurea cp genome revealed its conserved nature without gene loss or rearrangements when compared to other species of the Collabieae tribe. Phylogenetic analysis of Collabieae species also revealed that S. aurea has a distant evolutionary relationship to other members of the Collabieae species, despite the presence of problematic genera such as Phaius and Cephalantheropsis.

Evaluation and characteristic analysis of SSRs from the transcriptomic sequences of Perilla crop (Perilla frutescens L.)

Perilla crop is a self-fertilizing annual plant, cultivated and used mainly in East Asia. Perilla frutescens var. frutescens seeds are rich in unsaturated fatty acids, which have health benefits, and Perilla frutescens var. crispa leaves are rich in anthocyanins. However, genomic analysis such as whole genome sequencing or genetic mapping has not been performed on Perilla crop. This current study confirms the abundance and diversity of 15,991 simple sequence repeats (SSRs) classified in previous studies in the Perilla genome, selects and designs 1,538 SSR primer sets, and confirms which SSR primer sets exhibit high polymorphism. Of the 15,991 SSRs classified, there were 9,910 (62%) dinucleotide repeats, 5,652 (35.3%) trinucleotide repeats, and 429 (2.7%) tetranucleotide repeats. Among these, the most identified was (CT)n with a total of 4,817. The 15,991 SSRs had 4 to 26 repeats. Four repeats were the most frequent with 11,084 (69.3%). A total of 1,538 SSR primers were selected and designed to confirm polymorphism, of which 157 showed persistent and clear polymorphism. Among these 157 SSR primer sets, 98 (62.4%) were dinucleotide repeats, 39 (24.8%) were trinucleotide repeats, and 20 (12.7%) were tetranucleotide repeats. Among 549 SSR primers that showed polymorphism, trinucleotide repeats showed persistent polymorphism at a high rate. Therefore, when developing SSR primer sets for Perilla crop in the future, it is recommended that trinucleotide repeats be selected first. These research results will be helpful in future genomic analysis and development of SSR primers in Perilla crop.

Characterisation of Phytophthora capsici, causal agent of phytophthora foot rot disease of Piper nigrum L. in Sarawak, Malaysia

Phytophthora capsici causes Phytophthora foot rot disease on Piper nigrum. The pathogen lacks comprehensive study in Sarawak, Malaysia. In this study, P. capsici was isolated and characterised morphologically and molecularly from ten farms across three divisions in Sarawak. The pathogenicity of the isolates was confirmed, and distinct colony growth patterns, sporangia morphologies, and radial growth rates were observed across different media. Torulose hyphae structures were consistently observed, and compatible mating types were present. Fifty-two isolates retrieved from this study were genotyped using five Inter Simple Sequence Repeats and three Random Amplified Microsatellites primers. The UPGMA tree showed a close relationship between populations. A higher diversity within populations (73%) than among populations (27%) was observed. This study provides the first baseline data on the diversity of P. capsici in Sarawak, offering insights for managing the disease’s impact on pepper cultivation and the potential risk of increasing genetic diversity.

Land use to agriculture and planted forests strongly affect the genetic diversity of Baccharis crispa Spreng., a native herb of South America

Abstract Human population growth constantly requires an increase in the production of food and products from the timber industry. To meet this demand, agriculture and planted forests are advancing over natural areas. In view of this, it is necessary to know the effects of land use for different purposes (grain production, pastures, planted forests, fruit production, among other uses) on the genetic diversity of populations of native species. This knowledge can assist in land use planning as well as in the development of conservation strategies for native species. In this study, we evaluated the effect of land use for agriculture (mainly for cereal production) and planted forests on the genetic diversity of Baccharis crispa Spreng., a herb native to South America. To achieve our goals, we compared population genetic data obtained with three molecular markers (microsatellites, ISSR [inter-simple sequence repeat] and isoenzymes) with data on land use for agriculture and planted forests from 15 different locations. Our results showed that regardless of the molecular marker used, the greater the use of land for agriculture and planted forests, the lower was the genetic diversity of B. crispa populations. Baccharis crispa is a semi-perennial species that needs at least one year to reach its reproductive period, which is prevented in agricultural areas due to the land being turned over or dissected with herbicides every six months. In the studied regions, the planted forests are of eucalypt and/or pine, which besides being species with a high production of allelopathic substances, produce strong shading and B. crispa is a species that inhabits open grassland that needs high incidence of sunlight for development. The data obtained in our study can assist in the decision-making to use land in order to reconcile the production of supplies for humanity and for the conservation of nature.

Small striatal huntingtin inclusions in patients with motor neuron disease with reduced penetrance and intermediate HTT gene expansions.

Short tandem repeat expansions in the human genome are overrepresented in a variety of neurological disorders. It was recently shown that huntingtin (HTT) repeat expansions with full penetrance, i.e. 40 or more CAG repeats, which normally cause Huntington's disease (HD), are overrepresented in patients with amyotrophic lateral sclerosis (ALS). Whether patients carrying HTT repeat expansions with reduced penetrance, (36-39 CAG repeats), or alleles with intermediate penetrance, (27-35 CAG repeats), have an increased risk of ALS has not yet been investigated. Here, we examined the role of HTT repeat expansions in a motor neuron disease (MND) cohort, searched for expanded HTT alleles, and investigated correlations with phenotype and neuropathology. MND patients harboring C9ORF72 hexanucleotide repeat expansions (HREs) were included, to investigate whether HTT repeat expansions were more common in this group. We found a high prevalence of intermediate (range 5.63%-6.61%) and reduced penetrance (range 0.57%-0.66%) HTT gene expansions in this cohort compared to other populations of European ancestry, but no differences between the MND cohort and the control cohort were observed, regardless of C9ORF72HRE status. Upon autopsy of three patients with intermediate or reduced penetrance HTT alleles, huntingtin inclusions were observed in the caudate nucleus and frontal lobe, but no significant somatic mosaicism was detected in different parts of the nervous system. Thus, we demonstrate, for the first time, huntingtin inclusions in individuals with MND and intermediate and reduced penetrance HTT repeat expansions but more clinicopathological investigations are needed to further understand the impact of HTT gene expansion-related pleiotropy.

Comparative phylogenomic study of East Asian endemic genus, Corchoropsis Siebold & Zucc. (Malvaceae s.l.), based on complete plastome sequences

BackgroundEndemic plants are key to understanding the evolutionary history and enhancing biodiversity within their unique regions, while also offering significant economic potential. The East Asian endemic genus Corchoropsis Siebold & Zucc., classified within the subfamily Dombeyoideae of Malvaceae s.l., comprises three species.ResultsThis study characterizes the complete plastid genomes (plastomes) of C. crenata var. crenata Siebold & Zucc. and C. crenata var. hupehensis Pamp., which range from 160,093 to 160,724 bp. These genomes contain 78 plastid protein-coding genes, 30 tRNA, and four rRNA, except for one pseudogene, infA. A total of 316 molecular diagnostic characters (MDCs) specific to Corchoropsis were identified. In addition, 91 to 92 simple sequence repeats (SSRs) in C. crenata var. crenata and 75 in C. crenata var. hupehensis were found. Moreover, 49 long repeats were identified in both the Chinese C. crenata var. crenata and C. crenata var. hupehensis, while 52 were found in the South Korean C. crenata var. crenata. Our phylogenetic analyses, based on 78 plastid protein-coding genes, reveal nine subfamilies within the Malvaceae s.l. with high support values and confirm Corchoropsis as a member of Dombeyoideae. Molecular dating suggests that Corchoropsis originated in the Oligocene, and diverged during the Miocene, influenced by the climate shift at the Eocene–Oligocene boundary.ConclusionsThe research explores the evolutionary relationships between nine subfamilies within the Malvaceae s.l. family, specifically identifying the position of the Corchoropsis in the Dombeyoideae. Utilizing plastome sequences and fossil data, the study establishes that Corchoropsis first appeared during the Eocene and experienced further evolutionary divergence during the Miocene, paralleling the evolutionary patterns observed in other East Asian endemic species.

Habitat fragmentation strongly restricts gene flow in endangered ectomycorrhizal fungal populations: Evidence from Rhizopogon togasawarius, specific to Pseudotsuga japonica, across the entire distribution range.

Habitat fragmentation reduces gene flow, causing genetic differentiation and diversity loss in endangered species through genetic drift and inbreeding. However, the impact of habitat fragmentation on ectomycorrhizal (ECM) fungi remains unexplored, despite their critical roles in forest ecosystems. Here, we investigated the population genetic structure and the demographic history of Rhizopogon togasawarius, the ECM fungus specifically colonizing the host tree Pseudotsuga japonica, across its entire distribution range (>200 km). These two species are designated as endangered species on the IUCN Red List since they are found only in small, fragmented forests in Japan. We analysed 236 R. togasawarius individuals from five remaining populations across the Kii Peninsula and the Shikoku Island, separated by a sea channel. Simple sequence repeat analyses using 20 loci revealed strong genetic differentiation among populations (FST = 0.255), even significant in the nearest population pair separated by a distance of only 8 km (FST = 0.075), indicating extremely limited gene flow between populations. DIYABC-RF analyses implied that population divergence occurred approximately 6000 generations ago between the two regions, and nearly 1500 generations ago between the nearest populations within Shikoku Island, related to past climate events. Because of prolonged genetic isolation, significant inbreeding was confirmed in four of five populations, where effective population sizes became very small (Ne = 9.0-58.0). Although evaluation of extinction risks for microorganisms is challenging, our conservation genetic results indicated that habitat fragmentation increases extinction risk through population genetic mechanisms, and therefore should not be overlooked in biodiversity conservation efforts.

Comparative mitochondrial genomics of Terniopsis yongtaiensis in Malpighiales: structural, sequential, and phylogenetic perspectives

BackgroundTerniopsis yongtaiensis, a member of the Podostemaceae family, is an aquatic flowering plant displaying remarkable adaptive traits that enable survival in submerged, turbulent habitats. Despite the progressive expansion of chloroplast genomic information within this family, mitochondrial genome sequences have yet to be reported.ResultsIn current study, the mitochondrial genome of the T. yongtaiensis was characterized by a circular genome of 426,928 bp encoding 31 protein-coding genes (PCGs), 18 tRNAs, and 3 rRNA genes. Our comprehensive analysis focused on gene content, repeat sequences, RNA editing processes, intracellular gene transfer, phylogeny, and codon usage bias. Numerous repeat sequences were identified, including 130 simple sequence repeats, 22 tandem repeats, and 220 dispersed repeats. Phylogenetic analysis positioned T. yongtaiensis (Podostemaceae) within the Malpighiales order, showing a close relationship with the Calophyllaceae family, which was consistent with the APG IV classification. A comparative analysis with nine other Malpighiales species revealed both variable and conserved regions, providing insights into the genomic evolution within this order. Notably, the GC content of T. yongtaiensis was distinctively lower compared to other Malpighilales, primarily due to variations in non-coding regions and specific protein-coding genes, particularly the nad genes. Remarkably, the number of RNA editing sites was low (276), distributed unevenly across 27 PCGs. The dN/dS analysis showed only the ccmB gene of T. yongtaiensis was positively selected, which plays a crucial role in cytochrome c biosynthesis. Additionally, there were 13 gene-containing homologous regions between the mitochondrial and chloroplast genomes of T. yongtaiensis, suggesting the gene transfer events between these organellar genomes.ConclusionsThis study assembled and annotated the first mitochondrial genome of the Podostemaceae family. The comparison results of mitochondrial gene composition, GC content, and RNA editing sites provided novel insights into the adaptive traits and genetic reprogramming of this aquatic eudicot group and offered a foundation for future research on the genomic evolution and adaptive mechanisms of Podostemaceae and related plant families in the Malpighiales order.

Molecular fingerprinting of highly resistant maize lines to turcicum leaf blight

AbstractThe present study generates information related to the molecular divergence between turcicum leaf blight (TLB)-resistant and -susceptible lines. During molecular diversity studies, a total of 212 alleles were detected at 75 marker loci and ranged from two to five with an average of 2.83 alleles per locus. A direct correlation for the number of alleles and polymorphism information content (PIC) values was ascertained. For instance, marker phi123 produced high number of alleles (5) with PIC values of 0.77. Using the DARwin 6.0 programme, the UPGMA dendrogram grouped 40 maize inbreds into two distinct clusters, cluster-I (36 inbreds) and cluster-II (4 inbreds). Cluster-I contained two subclusters; the first subcluster contained 28 inbreds and the second subcluster contained eight inbreds whereas cluster-II contained four inbreds. This major cluster-II was further classified into two subclusters which contained two inbreds each. Most of the inbred lines except V-25 from cluster-II were highly resistant to TLB disease. These inbred lines can be used in crossing programmes to develop TLB-resistant hybrids by using divergent parents. In this study, allelic diversity and PIC values indicated a good efficiency of markers for studying the polymorphism level available in studied inbred lines. High level of diversity among the inbreds detected with simple sequence repeat markers indicated their suitability for the further breeding programme.

GENETIC DIVERSITY OF Moringa oleifera Lam. IN SOUTHEASTERN MEXICO

Moringa oleifera Lam. is a plant with nutritional and nutraceutical properties. It was introduced from India to Mexico, and some of its ecotypes are cultivated for industrial use. These ecotypes show morphological variations, indicating genetic variation. Therefore, the purpose of this study was to evaluate eight simple sequence repeat (SSR) loci to assess the genetic diversity of 10 cultivated M. oleifera populations collected from the southeast of Mexico. All markers generated a total of 127 alleles, with 15.8 alleles per locus. The polymorphic information content of markers was 0.869, with an average heterozygosity of 0.194. At the population level, the Nei´s gene diversity was found to be 0.66, with 4.9 different alleles per locus and 5.2 genotypes on average per accession. The distribution of genetic variation was 20 % among populations, 58 % between individuals, and 22 % within individuals. The principal coordinate analysis (PcoA) plot and unweighted pair group method with arithmetic mean (UPGMA) linkage revealed three groups, corresponding to the three original genetic populations identified through structure analysis (Delta K = 3). Results showed significant differences in genetic diversity in M. oleifera populations stablished in Southeastern Mexico, which can be leveraged to increase the use of this germplasm.