A Scalable and Robust Chloroplast Genotyping Solution: Development and Application of SNP and InDel Markers in the Maize Chloroplast Genome.

Maize(Zea mays. L) is a globally important crop, and understanding its genetic diversity is crucial for plant breeding phylogenetic analyses and comparative genetics. While nuclear markers have been extensively used for mapping agriculturally important genes, they are limited in recognizing characteristics, such as cytoplasmic male sterility and reciprocal cross hybrids. In this study, we performed next-generation sequencing of 176samples, and the maize cultivars represented five distinct groups. A total of 89 single nucleotide polymorphisms (SNPs) and 11 insertion/deletion polymorphisms (InDels) were identified. To enable high-throughput detection, we successfully amplified and confirmed 49 SNP and InDel markers, which were defined as a Varietal Chloroplast Panel (VCP) using the Kompetitive Allele Specific PCR (KASP). The specific markers provided a valuable tool for identifying chloroplast groups. The verification experiment, focusing on the identification of reciprocal cross hybrids and cytoplasmic male sterility hybrids, demonstrated the significant advantages of VCP markers in maternal inheritance characterization. Furthermore, only a small subset of these markers is needed to provide useful information, showcasing the effectiveness of these markers in elucidating the artificial selection process of elite maize lines.

Comparison of Clinical and Genetic Characteristics Between Younger and Older Lung Cancer Patients

Introduction The aim of this study was to analyze the clinical and genetic characteristics of young lung cancer cases, and to compare them with those of older cases. Methods We used the Thoracic Tumors Registry (TTR) as a data source representative of lung cancer cases diagnosed in Spain, and included all cases registered until 9/01/2023 which had information on age at diagnosis or the data needed to calculate it. We performed a descriptive statistical analysis and fitted logistic regressions to analyze how different characteristics influenced being a younger lung cancer patient. Results A total of 26,336 subjects were included. Lung cancer cases <50 years old had a higher probability of being women (OR: 1.38; 95% CI: 1.21–1.57), being in stage III or IV (OR: 1.32; 95% CI: 1.08–1.62), not having comorbidities (OR: 5.21; 95% CI: 4.59–5.91), presenting with symptoms at diagnosis (OR: 1.53; 95% CI: 1.29–1.81), and having ALK translocation (OR: 7.61; 95% CI: 1.25–46.32) and HER2 mutation (OR: 5.71; 95% CI: 1.34–24.33), compared with subjects ≥50 years. Among subjects <35 years old (n=61), our study observed a higher proportion of women (59.0% vs. 26.6%; p<0.001), never smokers (45.8% vs. 10.3%; p<0.001), no comorbidities (21.3% vs. 74.0%; p<0.001); ALK translocation (33.3% vs. 4.4%; p<0.001) and ROS1 mutation (14.3% vs. 2.3%; p=0.01), compared with subjects ≥35 years. Conclusions Lung cancer displays differences by age at diagnosis which may have important implications for its clinical management (AU)

Plant height variation and genetic diversity between Prunus ledebouriana (Schlecht.) YY Yao and Prunus tenella Batsch based on using SSR markers in East Kazakhstan.

Background: Genetic differences between isolated endemic populations of plant species and those with widely known twin species are relevant for conserving the biological diversity of our planet's flora. Prunus ledebouriana (Schlecht.) YY Yao is an endangered and endemic species of shrub almond from central Asia. Few studies have explored this species, which is closely related and morphologically similar to the well-known Prunus tenella Batsch. In this article, we present a comparative analysis of studies of three P. ledebouriana populations and one close population of P. tenella in Eastern Kazakhstan in order to determine the particular geographic mutual replacement of the two species. Methods: The populations were collected from different ecological niches, including one steppe population near Ust-Kamenogorsk (P. tenella) and three populations (P. ledebouriana) in the mountainous area. Estimation of plant height using a t-test suggested a statistically significant difference between the populations and the two species (P < 0.0001). DNA simple sequence repeat (SSR) markers were applied to study the two species' genetic diversity and population structure. Results: A total of 19 polymorphic SSR loci were analyzed, and the results showed that the population collected in mountainous areas had a lower variation level than steppe populations. The highest level of Nei's genetic diversity index was demonstrated in the 4-UK population (0.622) of P. tenella. The lowest was recorded in population 3-KA (0.461) of P. ledebouriana, collected at the highest altitude of the four populations (2,086 meters above sea level). The total genetic variation of P. ledebouriana was distributed 73% within populations and 27% between populations. STRUCTURE results showed that two morphologically similar species diverged starting at step K = 3, with limited population mixing. The results confirmed the morphological and genetic differences between P. tenella and P. ledebouriana and described the level of genetic variation for P. ledebouriana. The study's results proved that the steppe zone and mountain altitude factor between P. tenella and isolated mountain samples of P. ledebouriana.

Development of genetic markers in Yarrowia lipolytica.

The oleaginous yeast Yarrowia lipolytica represents a potential microbial cell factory for the recombinant production of various valuable products. Currently, the commonly used selection markers for transformation in Y. lipolytica are limited, and successive genetic manipulations are often restricted by the number of available selection markers. In our study, we developed a dominant marker, dsdA, which encodes a D-serine deaminase for genetic manipulation in Y. lipolytica. In Y. lipolytica, this marker confers the ability to use D-serine as a nitrogen source. In addition, the selection conditions of several infrequently used dominant markers including bleoR (zeocin resistance), kanMX (G418 resistance), and guaB (mycophenolic acid resistance) were also analyzed. Our results demonstrated that these selection markers can be used for the genetic manipulation of Y. lipolytica and their selection conditions were different for various strains. Ultimately, the selection markers tested here will be useful to expand the genetic toolbox of Y. lipolytica. KEY POINTS: • The dsdA from Escherichia coli was developed as a dominant marker. • The applicability of several resistance markers in Y. lipolytica was determined. • We introduced the Cre/mutant lox system for marker recycling.

Biomarkers Assessing the Role of Cumulus Cells on IVF Outcomes: A Systematic Review.

PURPOSE: Although significant improvements in assisted reproductive technology (ART) outcomes have been accomplished, a critical question remains: which embryo is most likely to result in a pregnancy? Embryo selection is currently based on morphological and genetic criteria; however, these criteria do not fully predict good-quality embryos and additional objective criteria are needed. The cumulus cells are critical for oocyte and embryo development. This systematic review assessed biomarkers in cumulus-oocyte complexes and their association with successful IVF outcomes. METHODS: A comprehensive search was conducted using PubMed, Embase, Scopus, and Web of Science from inception until November 2022. Only English-language publications were included. Inclusion criteria consisted of papers that evaluated genetic biomarkers associated with the cumulus cells (CCs) in humans and the following three outcomes of interest: oocyte quality, embryo quality, and clinical outcomes, including fertilization, implantation, pregnancy, and live birth rates. RESULTS: The search revealed 446 studies of which 42 met eligibility criteria. Nineteen studies correlated genetic and biochemical biomarkers in CCs with oocyte quality. A positive correlation was reported between oocyte quality and increased mRNA expression in CCs of genes encoding for calcium homeostasis (CAMK1D), glucose metabolism (PFKP), extracellular matrix (HAS2, VCAN), TGF-ß family (GDF9, BMP15), and prostaglandin synthesis (PTGS2). Nineteen studies correlated genetic and biochemical biomarkers in CCs with embryo quality. A positive correlation was reported between embryo quality and increased mRNA expression in CCs of genes encoding for extracellular matrix (HAS2), prostaglandin synthesis (PTGS2), steroidogenesis (GREM1), and decreased expression of gene encoding for hormone receptor (AMHR2). Twenty-two studies assessed genetic and biochemical biomarkers in CCs with clinical outcomes. Increased expression of genes encoding for extracellular matrix (VCAN), and TGF-ß family (GDF9, BMP15) were positively correlated with pregnancy rate. CONCLUSION: Genetic biomarkers from cumulus cells were associated with oocyte quality (CAMK1D, PFKP, HAS2, VCAN, GDF-9, BMP-15, PTGS2), embryo quality (GREM1, PTGS2, HAS2), and pregnancy rate (GDF9, BMP15, VCAN). These results might help guide future studies directed at tests of cumulus cells to devise objective criteria to predict IVF outcomes.

[Polymorphisms of OPRM1, OPRK1, DCC genes and non-suicidal self-injuries in adults]. / Polimorfizmy genov OPRM1, OPRK1 i DCC i nesuitsidal'nye samopovrezhdeniya u vzroslykh.

OBJECTIVE: To investigate the associations of OPRM1 gene rs179971, OPRK1 gene rs6473797 and DCC gene rs8084280 polymorphisms with non-suicidal self-injury (NSSI) characteristics and motivations in adults. MATERIAL AND METHODS: A pilot sample included 28 adult patients with history of NSSI (89.3% (n=25) women, median age (Q1-Q3) - 23 (21.25-25) years). Most patients (78.6%, n=20) had a diagnosis of bipolar disorder. NSSI characteristics and motivations were assessed using the Inventory of Statements about Self-Injury (ISAS) scale. The Childhood Trauma Questionnaire (CTQ) was used to control for childhood trauma - one of the most important environmental factors associated with NSSI. The Baratt Impulsivity Scale (BIS) and the Buss-Perry Aggression Questionnaire (BPAQ) were also used to assess impulsivity and aggression, respectively. RT-PCR was used for genotyping, a genetic effect was assessed using the dominant model. Mann-Whitney U-test, Pearson χ2-test and multiple linear regression were used for statistical analysis. RESULTS: Carriers of the minor G allele of OPRM1 gene rs1779971 had a higher level of aggression assessed by BPAQ (p=0.02). The minor C allele of OPRK1 gene rs6473797 was associated with an increase of the subjective importance of «Affect regulation¼ (B=2.23; CI 95% [0.39-4.06]; p=0.022) and «Anti-dissociation¼ (B=3.31; CI 95% [0.18-6.44]; p=0.039) motivations, whereas the minor T allele of DCC gene rs8084280, on the contrary, was associated with a decrease of the importance of «Affect regulation¼ (B=-1.74; CI 95% [-3.30 - -0.18]; p=0.032). Moreover, this effect was found after adjusting for diagnosis, sex, age, and the presence of childhood trauma. CONCLUSIONS: To our knowledge, this is the first study on the association of genetic markers with NSSI motivations. The results of this pilot study demonstrate that OPRK1 and DCC gene polymorphisms can determine differences in motivations for self-harm, however, these results require confirmation in large samples.

Multi-genome comprehensive identification of SSR/SV and development of molecular markers database to serve Sorghum bicolor (L.) breeding.

BACKGROUND: As an important food and cash crop, identification of DNA molecular markers is of great significance for molecular marker-assisted breeding of Sorghum (Sorghum bicolor (L.) moench). Although some sorghum-related mutation databases have been published, the special SSR and SV databases still need to be constructed and updated. RESULTS: In this study, the quality of 18 different sorghum genomes was evaluated, and two genomes were assembled at chromosome level. Through the identification and comparative analysis of SSR loci in these genomes, the distribution characteristics of SSR in the above sorghum genomes were initially revealed. At the same time, five representative reference genomes were selected to identify the structural variation of sorghum. Finally, a convenient SSR/SV database of sorghum was constructed by integrating the above results ( http://www.sorghum.top:8079/ ; http://43.154.129.150:8079/ ; http://47.106.184.91:8079/ ). Users can query the information of related sites and primer pairs. CONCLUSIONS: Anyway, our research provides convenience for sorghum researchers and will play an active role in sorghum molecular marker-assisted breeding.

Genetic Markers Regulating Blood Pressure in Extreme Discordant Sib Pairs.

Genome-wide scans performed in affected sib pairs have revealed small and often inconsistent clues to the loci responsible for the inherited components of hypertension. Since blood pressure is a quantitative trait regulated by many loci, two siblings at opposite extremes of the blood pressure distribution are more likely to have inherited different alleles at any given locus. Hence, we investigated an extreme discordant sib pair strategy to analyse markers from two previous loci of interest: (1) the Gordons syndrome locus that includes the WNK4 gene and (2) the ROMK locus identified in our first genome-wide scan. For this study, 24 sib pairs with strong family histories of essential hypertension were selected from the top and bottom 10% of the blood pressure distribution and genotyped for highly polymorphic microsatellite markers on chromosomes 11 and 17. The mean age of the population was 39.8 ± 7.8 years. A significant inverse correlation was found between the squared difference in pulse pressure and the number of alleles shared by IBD between the siblings for the DS11925 marker (r = -0.44, p = 0.031), systolic pressure and chromosome 17 markers (D17S250: r = -0.42, p = 0.040; D17S799 (r = -0.51, p = 0.011), and this relationship persisted after correcting for age and gender. Markers on chromosome 17 (D17S250, D17S928 and D17S1301) and 11 (D11S1999) also correlated with diastolic pressure. These results illustrate the successful use of discordant sib pair analysis to detect linkage within relatively small numbers of pedigrees with hypertension. Further analysis of this cohort may be valuable in complementing findings from the large genome wide scans in affected sib pairs.

Development of a molecular marker for the Pi1 gene based on the association of the SNAP protocol with the touch-up gradient amplification method.

Genetic resistance is the most effective and eco-friendly approach to combat rice blast. The application of resistance genes may be facilitated by the availability of molecular markers that allow marker-assisted selection during the breeding process. The Pi1 gene, considered to be a broad-spectrum resistance gene, might contribute to enhancing resistance to rice blast, but it lacks a suitable marker that can be used. In this study, we investigated nucleotide polymorphism in the Pik locus and combined the SNAP protocol with the touch-up gradient amplification method to develop a SNAP marker. The Pi1 SNAP marker could distinguish Pi1 from Pik alleles, and when used for screening a germplasm bank and an F2 population, it consistently identified germplasms carrying the Pi1 gene. The P1 SNAP marker offers as advantages to involve only the presence/absence analysis of PCR amplicons resolved on an agarose gel.

Discrimination of Camellia cultivars using iD-NA analysis.

Recently, many new cultivars have been taken abroad illegally, which is now considered an international issue. Botanical evidence found at a crime scene provides valuable information about the origin of the sample. However, botanical resources for forensic evidence remain underutilized because molecular markers, such as microsatellites, are not available without a limited set of species. Multiplexed intersimple sequence repeat (ISSR) genotyping by sequencing (MIG-seq) and its analysis method, identification of not applicable (iD-NA), have been used to determine several genome-wide genetic markers, making them applicable to all plant species, including those with limited available genetic information. Camellia cultivars are popular worldwide and are often planted in many gardens and bred to make new cultivars. In this study, we aimed to analyze Camellia cultivars/species through MIG-seq. MIG-seq could discriminate similar samples, such as bud mutants and closely related samples that could not be distinguished based on morphological features. This discrimination was consistent with that of a previous study that classified cultivars based on short tandem repeat (STR) markers, indicating that MIG-seq has the same or higher discrimination ability as STR markers. Furthermore, we observed unknown phylogenetic relationships. Because MIG-seq can be applied to unlimited species and low-quality DNA, it may be useful in various scientific fields.

Comparative Assessment of SSR and RAPD markers for genetic diversity in some Mango cultivars.

Genetic improvement mainly depends on the level of genetic variability present in the population, and the degree of genetic diversity in a population largely determines the rate of genetic advancement. For analyzing genetic diversity and determining cultivar identities, a molecular marker is a useful tool. Using 30 SSR (simple sequence repeat) and 30 RAPD (randomly amplified polymorphic DNA) markers, this study evaluated the genetic divergence of 17 mango cultivars. The effectiveness of the two marker systems was evaluated using their genetic diversity characteristics. Additionally, the effects of SM (simple matching) and Dice similarity coefficients and their effects on mango clustering were evaluated. The findings showed that SSR markers generated 192 alleles, all of which were polymorphic (100%). With RAPD markers, 434 bands were obtained, 361 of which were polymorphic (83%). The average polymorphic information content (PIC) for RAPD and SSR was 0.378 and 0.735, respectively. Using SSR markers resulted in much higher values for other genetic diversity parameters compared to RAPD markers. Furthermore, grouping the genotypes according to the two similarity coefficients without detailed consideration of these coefficients could not influence the study results. The RAPD markers OPA_01, OPM_12 followed by OPO_12 and SSR markers MIAC_4, MIAC_5 followed by mMiCIR_21 were the most informative in terms of describing genetic variability among the cultivars under study; they can be used in further investigations such as genetic mapping or marker-assisted selection. Overall, 'Zebda' cultivar was the most diverse of the studied cultivars.

Mexican Biobank advances population and medical genomics of diverse ancestries.

Latin America continues to be severely underrepresented in genomics research, and fine-scale genetic histories and complex trait architectures remain hidden owing to insufficient data1. To fill this gap, the Mexican Biobank project genotyped 6,057 individuals from 898 rural and urban localities across all 32 states in Mexico at a resolution of 1.8 million genome-wide markers with linked complex trait and disease information creating a valuable nationwide genotype-phenotype database. Here, using ancestry deconvolution and inference of identity-by-descent segments, we inferred ancestral population sizes across Mesoamerican regions over time, unravelling Indigenous, colonial and postcolonial demographic dynamics2-6. We observed variation in runs of homozygosity among genomic regions with different ancestries reflecting distinct demographic histories and, in turn, different distributions of rare deleterious variants. We conducted genome-wide association studies (GWAS) for 22 complex traits and found that several traits are better predicted using the Mexican Biobank GWAS compared to the UK Biobank GWAS7,8. We identified genetic and environmental factors associating with trait variation, such as the length of the genome in runs of homozygosity as a predictor for body mass index, triglycerides, glucose and height. This study provides insights into the genetic histories of individuals in Mexico and dissects their complex trait architectures, both crucial for making precision and preventive medicine initiatives accessible worldwide.

Pyramiding resistance genes for bacterial leaf blight (Xanthomonas oryzae pv. Oryzae) into the popular rice variety, Pratikshya through marker assisted backcrossing.

BACKGROUND: Bacterial leaf blight (BLB) is one of the major biotic stress in rice cultivation. Management techniques, such as the development of BLB-resistant cultivars, are required to lessen the severity of the disease attack and yield losses. Pratikshya was selected in the present investigation as the recipient parent, as it is one of the popular high-yielding rice varieties of Odisha, India, which is having excellent grain as well as cooking quality. However, Pratikshya is highly susceptible to BLB which is prevalent in Eastern Indian region. METHODS AND RESULTS: Three major BLB resistance genes xa5, xa13, and Xa21 from the donor source Swarna MAS (CR Dhan 800) were attempted to introduce into Pratikshya through a marker-assisted backcross breeding program. Those markers closely linked to the target genes were employed for foreground selection in the segregating generations till BC2F3. In each backcross generation, progenies containing all three targeted resistance genes and phenotypically more similar to the recipient parent, Pratikshya were selected and backcrossed. Screening of 1,598 plants of the BC2F2 population was conducted against BLB using Xoo inoculum and 35 resistant plants similar to Pratikshya were carried forward to the next generation. In the BC2F3 generation, 31 plants were found to possess all the three resistance genes. For background selection of plants carrying resistance genes 45 polymorphic SSR markers were employed. Evaluation of the pyramided lines at BC2F4 generation exhibited that, most pyramided lines were similar to Pratikshya in terms of morphological features and yield parameters, and some lines were superior to the recurrent parent in terms of morphological features and yield parameters. CONCLUSION: The three-gene pyramided lines showed a high level of resistance to BLB infection and are anticipated to offer a significant yield advantage over the recipient parent Pratikshya. The pyramided lines can further be used for multi-location trial, so as to be released as a variety or can be used as a potential donor for BLB resistance genes.

DNA Markers, Pathogenicity Test, and Multilocus Sequence Analysis to Differentiate and Characterize Cereal-Specific Xanthomonas translucens Strains.

Xanthomonas translucens contains a group of bacterial pathogens that are closely related and have been divided into several pathovars based on their host range. X. translucens pv. undulosa (Xtu) and X. translucens pv. translucens (Xtt) are two important pathovars that cause bacterial leaf streak disease on wheat and barley, respectively. In this study, DNA markers were developed to differentiate Xtu and Xtt and were then used to characterize a collection of X. translucens strains with diverse origins, followed by confirmation and characterization with pathogenicity tests and multilocus sequence analysis/typing (MLSA/MLST). We first developed cleaved amplified polymorphic sequence markers based on the single-nucleotide polymorphisms within a cereal pathovar-specific DNA sequence. In addition, two Xtt-specific markers, designated Xtt-XopM and Xtt-SP1, were developed from comparative genomics among the sequenced Xtt/Xtu genomes. Using the developed markers, a collection of X. translucens strains were successfully identified as Xtu or Xtt. Pathogenicity tests on wheat and barley plants and MLSA of four housekeeping genes validated the pathovar assignation of those strains. Furthermore, MLSA revealed distinct subclades within both Xtu and Xtt groups. Seven and three sequence types were identified from MLST for Xtu and Xtt strains, respectively. The establishment of efficient Xtt/Xtu differentiation methods and characterization of those strains will be useful in studying disease epidemiology and host-pathogen interactions and breeding programs when screening for sources of resistance for these two important bacterial pathogens.

Refinement of Rf1-gene localization and development of the new molecular markers for fertility restoration in sunflower.

BACKGROUND: Ability to restore male fertility is important trait for sunflower breeding. The most commonly used fertility restoration gene in the production of sunflower hybrids is Rf1. The localization of Rf1 on the linkage group 13 has been previously shown, however, its exact position, its sequence and molecular mechanism for fertility restoration remain unknown. Therefore, several markers linked to Rf1 gene, commonly used for MAS, don't always allow to identify the genotype of plants. For this reason, the search for new markers and precise localization of the Rf1 gene is an urgent task. METHODS AND RESULTS: Based on previously identified single nucleotide polymorphisms (SNPs) at LG13, significantly associated with the ability to restore male fertility, two markers have been developed that have performed well after careful evaluation. These markers, together with other Rf1 markers, were applied for genotyping 72 diversity panel accessions and 291 individuals of F2 segregating population, obtained from crossing the cytoplasmic male sterility (CMS) AHO33 and restorer RT085HO lines. The analysis revealed no recombinants between Rf1 gene and SRF833 marker, the distance between Rf1 and SRF122 marker was 1.0 cM. CONCLUSIONS: Data obtained made it possible to specify the localization of the Rf1 gene and reduce the list of candidate genes to the 3 closely linked PPR-genes spanning a total of 59 Kb. However, it cannot be ruled out that analysis of the candidate region in the genome of fertility restorer lines can reveal new candidate genes in this locus that are absent in the cytoplasmic male sterility maintainer reference sequence.

Genetic diversity of the breeding collection of tomato varieties in Kazakhstan assessed using SSR, SCAR and CAPS markers.

Tomato is one of the most prominent crops in global horticulture and an important vegetable crop in Kazakhstan. The lack of data on the genetic background of local varieties limits the development of tomato breeding in the country. This study aimed to perform an initial evaluation of the breeding collection of tomato varieties from the point of view of their genetic structure and pathogen resistance using a set of PCR based molecular markers, including 13 SSR markers for genetic structure analysis, and 14 SCAR and CAPS markers associated with resistance to five pathogens: three viruses, fungus Fusarium oxysporum, and oomycete P hytophthora infestans. Nine SSR markers were with a PIC value varying from 0.0562 (low information content) to 0.629 (high information content). A weak genetic structure was revealed in the samples of varieties including local cultivars and, predominantly, varieties from Russia and other ex-USSR countries. The local varieties were closely related to several groups of cultivars of Russian origin. Screening for a set of resistance markers revealed the common occurrence of the resistance locus I against Fusarium oxysporum and only the occasional presence of resistance alleles of other markers. No markers of resistance to the three considered viruses were revealed in local tomato varieties. Only two local cultivars had markers of resistance to P. infestans, and only the 'Meruert' cultivar had a combination of resistance markers against P. infestans and F. oxysporum. The obtained results have demonstrated the need for further studies of local tomato varieties with a wider range of molecular markers and source germplasm to lay a foundation for the development of tomato breeding in Kazakhstan.

Development of EST-SSRs based on the transcriptome of Castanopsis carlesii and cross-species transferability in other Castanopsis species.

Castanopsis carlesii (Hemsl.) Hay. is a widely distributed and dominant tree species native to subtropical China with significant ecological and economic value. Due to serious human-related disturbance, its wild resources have been increasingly reduced, and whether may result in the loss of genetic diversity. However, no population genetics studies of natural C. carlesii have been reported to date. Microsatellite markers have been a useful tool in population genetics. Therefore, we developed EST-SSR markers based on the transcriptome sequencing of C. carlesii leaves. A total of 149,380,224 clean reads were obtained, and 63,012 nonredundant unigenes with a mean length of 1,034 bp were assembled and annotated based on sequence similarity searches in the Nr, Nt, KO, SwissProt, PFAM, KOG, and GO databases. The results showed that only 5,559 (8.82%) unigenes were annotated in all seven databases, but 46,338 (73.53%) could be annotated in at least one database. A total of 31,459 potential EST-SSRs were identified in 18,690 unigenes, with an average frequency of one SSR approximately 2 kb. Among the 100 EST-SSR primer pairs designed, 49 primer pairs successfully produced the expected product by amplification, with a success rate of 49%, but only 20 primer pairs showed abundant polymorphisms. Polymorphisms were verified using 25 samples from C. carlesii in Qimen, Anhui. A total of 119 alleles were detected, with a mean number of alleles (Na) of 5.95 per locus and a mean polymorphism information content (PIC) of 0.6125. All the 20 newly developed EST-SSR markers were verified in other Castanopsis species (C. sclerophylla, C. lamontii, C. fargesii, C. eyrei and C. jucunda). Sixteen primer pairs showed successful amplification in all five Castanopsis species (80%), and the transferability ratios ranged from 90% to 100%. These developed EST-SSR markers can be applied to population genetic and germplasm evaluations of C. carlesii and related species.

Genetic Mapping of a Novel Gene PmAege7M from Aegilops geniculata Conferring Resistance to Wheat Powdery Mildew.

Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most damaging foliage diseases of wheat across the world. Aegilops geniculata Roth is a valuable gene resource for enhancing wheat resistance to powdery mildew. This study identified Ae. geniculata accession PI 487224 as immune and PI 487228 as susceptible to powdery mildew. Genetic analysis of the F1, F2, and F2:3 progeny derived from PI 487224 × PI 487228 showed that powdery mildew resistance in PI 487224 was controlled by two independent dominant genes located on two different nonhomologous chromosomes. By combing bulked segregant RNA-Seq, genetic linkage analysis of a single resistance gene segregation population, and marker analysis of a set of 14 wheat-Ae. geniculata chromosome addition lines, one of the resistance genes, temperately designated PmAege7M, was mapped to a 4.9-cM interval flanked by markers STS7-55926 and SNP7-45792/STS7-65911 on the long arm of chromosome 7 Mg of PI 487224, spanning 604.73 to 622.82 Mb on the 7D long arm based on the Ae. tauschii reference genome (Aet_v4.0). The map and closely linked markers of PmAege7M from Ae. geniculata in this study will facilitate the transfer of PmAege7M into common wheat and fine mapping of the gene.

Morpho-biochemical characterization and molecular marker based genetic diversity of pearl millet (Pennisetum glaucum (L.) R. Br.).

Pearl millet is a key food for millions living in semi-arid and arid regions and is a main diet for poorer populations. The genetic diversity existing in the pearl millet germplasm can be used to improve the micronutrient content and grain yield. Effective and organized exploitation of diversity at morphological and DNA levels is the strategy for any crop improvement program. In this study, the genetic diversity of 48 pearl millet genotypes was evaluated for eight morphological traits and eleven biochemical characters. All genotypes were also characterized using twelve SSR and six SRAP markers to evaluate genetic diversity. The significant mean difference between morphological and biochemical traits were detected. The productive tillers per plant varied from 2.65 to 7.60 with a mean of 4.80. The grain yield of genotypes varied more than 3× from 15.85 g (ICMR 07222) to 56.75 g (Nandi 75) with an average of 29.54 g per plant. Higher levels of protein, iron, and zinc contents were found to be present in ICMR 12555 (20.6%), ICMR 08666 (77.38 ppm), and IC 139900 (55.48 ppm), respectively, during the experiment. Substantial variability was observed for grain calcium as it ranged from 100.00 ppm (ICMR 10222) to 256.00 ppm (ICMR 12888). The top eight nutrient-dense genotypes flowered in 34-74 days and had 5.71-9.39 g 1,000 grain weight. Genotype ICMR 08666 was superior for Fe, Zn, K and P. The inter-genotype similarity coefficient at the genetic level, generated using DNA markers, ranged from 0.616 to 0.877 with a mean of 0.743. A combination of morpho-biochemical traits and DNA markers based diversity may help to differentiate the genotypes and diverse genotypes can be used in breeding programs to improve the mineral content in pearl millet.

Genome-wide discovery of di-nucleotide SSR markers based on whole genome re-sequencing data of Cicer arietinum L. and Cicer reticulatum Ladiz.

Simple sequence repeats (SSRs) are valuable genetic markers due to their co-dominant inheritance, multi-allelic and reproducible nature. They have been largely used for exploiting genetic architecture of plant germplasms, phylogenetic analysis, and mapping studies. Among the SSRs, di-nucleotide repeats are the most frequent of the simple repeats distributed throughout the plant genomes. In present study, we aimed to discover and develop di-nucleotide SSR markers by using the whole genome re-sequencing (WGRS) data from Cicer arietinum L. and C. reticulatum Ladiz. A total of 35,329 InDels were obtained in C. arietinum, whereas 44,331 InDels in C. reticulatum. 3387 InDels with 2 bp length were detected in C. arietinum, there were 4704 in C. reticulatum. Among 8091 InDels, 58 di-nucleotide regions that were polymorphic between two species were selected and used for validation. We tested primers for evaluation of genetic diversity in 30 chickpea genotypes including C. arietinum, C. reticulatum, C. echinospermum P.H. Davis, C. anatolicum Alef., C. canariense A. Santos & G.P. Lewis, C. microphyllum Benth., C. multijugum Maesen, C. oxyodon Boiss. & Hohen. and C. songaricum Steph ex DC. A total of 244 alleles were obtained for 58 SSR markers giving an average of 2.36 alleles per locus. The observed heterozygosity was 0.08 while the expected heterozygosity was 0.345. Polymorphism information content was found to be 0.73 across all loci. Phylogenetic tree and principal coordinate analysis clearly divided the accessions into four groups. The SSR markers were also evaluated in 30 genotypes of a RIL population obtained from an interspecific cross between C. arietinum and C. reticulatum. Chi-square (χ2) test revealed an expected 1:1 segregation ratio in the population. These results demonstrated the success of SSR identification and marker development for chickpea with the use of WGRS data. The newly developed 58 SSR markers are expected to be useful for chickpea breeders.