Evaluation of yellow rust resistance in backcross populations of wheat

Aim: To screen wheat populations derived from cross DBW17 × WH1105 for loci imparting yellow rust resistance and selection of plants using polymorphic SSRs. Methodology: The study for yellow rust resistance was carried out on two populations, i.e., BC1F2 and BC2F2. Stress was provided by planting infector rows between the blocks and by artificial inoculation using a mixture of races 46S102, 47S103 and 78S84 of stripe rust pathogen. DNA isolated from young leaves was checked for the presence of yellow rust resistance genes using gene specific primers. Results: Fifteen primers were found to be polymorphic among parents DBW17 and WH1105. Fifteen polymorphic SSR markers were dispersed over the wheat genome (AABBDD), with allele range 2-5. These polymorphic SSR markers were used to produce molecular diversity among progeny lines. Cluster analysis of parents and both the populations, showed that two parents were diverse genetically and in both backcrosses progeny lines resembled their respective recurrent parent. Single marker analysis using data revealed that primers on nine chromosomes were associated with grain yield per plant, other yield attributes and yellow rust resistance in both populations. Interpretation: This study showed that a linked marker like Xgwm582 could be a promising tool for breeding wheat with enhanced tolerance to yellow rust resistance. However, growth rates and biomass production provide reliable criteria for assessing the degree of yellow rust resistance and the ability of a plant to withstand it.

Identification of Candidate Genes and Genomic Regions Associated with Adult Plant Resistance to Stripe Rust in Spring Wheat

Wheat stripe rust (caused by Puccinia striiformis f. sp. tritici) is a major disease that damages wheat plants and affects wheat yield all over the world. In recent years, stripe rust became a major problem that affects wheat yield in Egypt. New races appeared and caused breakdowns in the resistant genotypes. To improve resistance in the Egyptian genotypes, new sources of resistance are urgently needed. In the recent research, a set of 95 wheat genotypes collected from 19 countries, including Egypt, were evaluated for their resistance against the Egyptian race(s) of stripe rust under field conditions in the two growing seasons 2018/2019 and 2019/2020. A high genetic variation was found among the tested genotypes. Single marker analysis was conducted using a subset of 71 genotypes and 424 diversity array technology (DArT) markers, well distributed across the genome. Out of the tested markers, 13 stable markers were identified that were significantly associated with resistance in both years (p-value ≤ 0.05). By using the sequence of the DArT markers, the chromosomal position of the significant DArT markers was detected, and nearby gene models were identified. Two markers on chromosomes 5A and 5B were found to be located within gene models functionally annotated with disease resistance in plants. These two markers could be used in marker-assisted selection for stripe rust resistance under Egyptian conditions. Two German genotypes were carrying the targeted allele of all the significant DArT markers associated with stripe rust resistance and could be used to improve resistance under Egyptian conditions.

Environmental and Genetic Factors Affecting Apospory Expressivity in Diploid Paspalum rufum

In angiosperms, gametophytic apomixis (clonal reproduction through seeds) is strongly associated with polyploidy and hybridization. The trait is facultative and its expressivity is highly variable between genotypes. Here, we used an F1 progeny derived from diploid apomictic (aposporic) genotypes of Paspalum rufum and two F2 families, derived from F1 hybrids with different apospory expressivity (%AES), to analyze the influence of the environment and the transgenerational transmission of the trait. In addition, AFLP markers were developed in the F1 population to identify genomic regions associated with the %AES. Cytoembryological analyses showed that the %AES was significantly influenced by different environments, but remained stable across the years. F1 and F2 progenies showed a wide range of %AES variation, but most hybrids were not significantly different from the parental genotypes. Maternal and paternal genetic linkage maps were built covering the ten expected linkage groups (LG). A single-marker analysis detected at least one region of 5.7 cM on LG3 that was significantly associated with apospory expressivity. Our results underline the importance of environmental influence in modulating apospory expressivity and identified a genomic region associated with apospory expressivity at the diploid level.

Genetic diversity may help evolutionary rescue in a clonal endemic plant species of Western Himalaya

Habitat loss due to climate change may cause the extinction of the clonal species with a limited distribution range. Thus, determining the genetic diversity required for adaptability by these species in sensitive ecosystems can help infer the chances of their survival and spread in changing climate. We studied the genetic diversity and population structure of Sambucus wightiana—a clonal endemic plant species of the Himalayan region for understanding its possible survival chances in anticipated climate change. Eight polymorphic microsatellite markers were used to study the allelic/genetic diversity and population structure. In addition, ITS1–ITS4 Sanger sequencing was used for phylogeny and SNP detection. A total number of 73 alleles were scored for 37 genotypes at 17 loci for 8 SSRs markers. The population structural analysis using the SSR marker data led to identifying two sub-populations in our collection of 37 S. wightiana genotypes, with 11 genotypes having mixed ancestry. The ITS sequence data show a specific allele in higher frequency in a particular sub-population, indicating variation in different S. wightiana accessions at the sequence level. The genotypic data of SSR markers and trait data of 11 traits of S. wightiana, when analyzed together, revealed five significant Marker-Trait Associations (MTAs) through Single Marker Analysis (SMA) or regression analysis. Most of the SSR markers were found to be associated with more than one trait, indicating the usefulness of these markers for working out marker-trait associations. Moderate to high genetic diversity observed in the present study may provide insurance against climate change to S. wightiana and help its further spread.

Genotyping-by-Sequencing Based Genetic Mapping Identified Major and Consistent Genomic Regions for Productivity and Quality Traits in Peanut

With an objective of identifying the genomic regions for productivity and quality traits in peanut, a recombinant inbred line (RIL) population developed from an elite variety, TMV 2 and its ethyl methane sulfonate (EMS)-derived mutant was phenotyped over six seasons and genotyped with genotyping-by-sequencing (GBS), Arachis hypogaea transposable element (AhTE) and simple sequence repeats (SSR) markers. The genetic map with 700 markers spanning 2,438.1 cM was employed for quantitative trait loci (QTL) analysis which identified a total of 47 main-effect QTLs for the productivity and oil quality traits with the phenotypic variance explained (PVE) of 10–52% over the seasons. A common QTL region (46.7–50.1 cM) on Ah02 was identified for the multiple traits, such as a number of pods per plant (NPPP), pod weight per plant (PWPP), shelling percentage (SP), and test weight (TW). Similarly, a QTL (7.1–18.0 cM) on Ah16 was identified for both SP and protein content (PC). Epistatic QTL (epiQTL) analysis revealed intra- and inter-chromosomal interactions for the main-effect QTLs and other genomic regions governing these productivity traits. The markers identified by a single marker analysis (SMA) mapped to the QTL regions for most of the traits. Among the five potential candidate genes identified for PC, SP and oil quality, two genes (Arahy.7A57YA and Arahy.CH9B83) were affected by AhMITE1 transposition, and three genes (Arahy.J5SZ1I, Arahy.MZJT69, and Arahy.X7PJ8H) involved functional single nucleotide polymorphisms (SNPs). With major and consistent effects, the genomic regions, candidate genes, and the associated markers identified in this study would provide an opportunity for gene cloning and genomics-assisted breeding for increasing the productivity and enhancing the quality of peanut.

Mapping QTLs controlling the biosynthesis of maltose in soybean

The present study was carried out to identify genomic regions associated with maltose in 2 F2 populations through assessment of sugars using HPLC and genotyping using SSR markers across the genome. SSR markers, Sat_216 (chr 12) and Satt681 (chr 6) in F2 population I and Sat_105 (chr 20) in F2 population II showed significant (P< 0.5) association with maltose content through single marker analysis (SMA) with LOD score of 3.18 (R2 =9.7), 2.54 (R2 =6.8), and 3.54 (R2 =10.4), respectively. Composite interval mapping analysis (CIM) let to identify different QTLs (other than SMA) for maltose content on chr 11, chr 13 and chr 17 in F2 population I while chr 6 and chr15 in F2 population II. QTLs identified for maltose content are in proximity of known functional genes responsible for degradation of starch into maltose. QTLs identified for maltose in the study may be deployed for improving efficiency of marker assisted breeding for development of soybean genotypes with high levels of this sugar.

Quantitative trait loci and candidate genes associated with freezing tolerance of winter triticale (× Triticosecale Wittmack)

Freezing tolerance of triticale is a major trait contributing to its winter hardiness. The identification of genomic regions — quantitative trait loci (QTL) and molecular markers associated with freezing tolerance in winter hexaploid triticale — was the aim of this study. For that purpose, a new genetic linkage map was developed for the population of 92 doubled haploid lines derived from ‘Hewo’ × ‘Magnat’ F1 hybrid. Those lines, together with parents were subjected to freezing tolerance test three times during two winter seasons. Plants were grown and cold-hardened under natural fall/winter conditions and then subjected to freezing in controlled conditions. Freezing tolerance was assessed as the plants recovery (REC), the electrolyte leakage (EL) from leaves and chlorophyll fluorescence parameters (JIP) after freezing. Three consistent QTL for several fluorescence parameters, electrolyte leakage, and the percentage of the survived plants were identified with composite interval mapping (CIM) and single marker analysis (SMA). The first locus Qfr.hm-7A.1 explained 9% of variation of both electrolyte leakage and plants recovery after freezing. Two QTL explaining up to 12% of variation in plants recovery and shared by selected chlorophyll fluorescence parameters were found on 4R and 5R chromosomes. Finally, main locus Qchl.hm-5A.1 was detected for chlorophyll fluorescence parameters that explained up to 19.6% of phenotypic variation. The co-located QTL on chromosomes 7A.1, 4R and 5R, clearly indicated physiological and genetic relationship of the plant survival after freezing with the ability to maintain optimal photochemical activity of the photosystem II and preservation of the cell membranes integrity. The genes located in silico within the identified QTL include those encoding BTR1-like protein, transmembrane helix proteins like potassium channel, and phosphoric ester hydrolase involved in response to osmotic stress as well as proteins involved in the regulation of the gene expression, chloroplast RNA processing, and pyrimidine salvage pathway. Additionally, our results confirm that the JIP test is a valuable tool to evaluate freezing tolerance of triticale under unstable winter environments.

​Inheritance Studies and Validation of Molecular Markers Associated with Botrytis Grey Mould in Chickpea (Cicer arietinum L.)

Background: The Botrytis grey mould (BGM) is a devastating foliar disease of chickpea. In order to develop resistant high yielding varieties, the genetic mechanism governing the inheritance of resistance against BGM must be decoded. The molecular markers associated with BGM resistance are also need to be validated for marker assisted pyramiding of BGM resistance in chickpea. Methods: The present study was conducted during rabi seasons of 2014-18 at GBPUAT, Pantnagar. The experimental material consisted of six generations (P1, P2, F1, F2, BC1 and BC2) of a cross between a BGM resistant variety GL 10006 and susceptible variety H 208. The disease data was scored on nine-point (1-9) scale and were subjected to chi-square analysis. The inheritance was also studied by using 28 STMS markers at the Pulse Breeding Laboratory of GBPUAT, Pantnagar during 2017-18. The Single Marker Analysis was performed for validation of markers associated with BGM. Result: The results indicated that resistance for BGM in chickpea is dominant over susceptibility. A major QTL i.e. TA118 along with some minor QTLs are involved in governing resistance to BGM. The marker TA118, TS72 and TA144 can be used effectively in marker assisted selection for getting desirable recombinants in chickpea breeding.

New PCR-specific markers for pollen fertility restoration QRfp-4R in rye (Secale cereale L.) with Pampa sterilizing cytoplasm

Pampa cytoplasmic male sterility phenomenon is used extensively in the rye hybrid breeding programs. It relies on sterilizing action of the cytoplasm resulting in non-viable pollen of female lines. The sterilizing effect is problematic for reversion, and efficient restores are needed. The most promising QTL is located on chromosome 4R, but other chromosomes may also code the trait. Advanced recombinant inbred lines formed bi-parental mapping population genotyped with DArTseq markers. Genetic mapping allowed the seven linkage groups to construct with numerous markers and represent all rye chromosomes. Single marker analysis and composite interval mapping were conducted to identify markers linked to the pollen fertility. Association mapping was used to detect additional markers associated with the trait. A highly significant QTL (QRfp-4R) that explained 42.3% of the phenotypic variation was mapped to the distal part of the long arm of the 4R chromosome. The markers localized in the QRfp-4R region achieve R2 association values up to 0.59. The homology of the 43 marker sequences to the loci responsible for fertility restoration in other species and transcription termination factor (mTERF) linked to Rf genes was established. Ten markers were successfully converted into PCR-specific conditions, and their segregation pattern was identical to that of unconverted DArTs.

Quantitative Trait Loci and Candidate Genes associated with Freezing Tolerance of Winter Triticale (×Triticosecale Wittmack)

Freezing tolerance of triticale is a major trait contributing to its winter hardiness. The identification of genomic regions – quantitative trait loci (QTLs) and molecular markers associated with freezing tolerance in winter hexaploid triticale was the aim of this study. For that purpose a new genetic linkage map was developed for the population of 92 doubled haploid lines derived from ‘Hewo’ × ‘Magnat’ F1 hybrid. Those lines, together with parents were subjected to freezing tolerance test three times during two winter seasons. Plants were grown and cold-hardened under natural fall/winter conditions and then subjected to freezing in controlled conditions. Freezing tolerance was assessed as the plants recovery (REC), the electrolyte leakage (EL) and chlorophyll fluorescence parameters (JIP) after freezing. Three consistent QTLs for several fluorescence parameters, electrolyte leakage and the percentage of the survived plants were identified with composite interval mapping (CIM) and single marker analysis (SMA). The first locus Qfr.hm-7A.1 explained 9 % of variation of both electrolyte leakage and plants recovery after freezing. Two QTLs explaining up to 12 % of variation in plants recovery and shared by selected chlorophyll fluorescence parameters were found on 4R and 5R chromosomes. Finally, main locus Qchl.hm-5A.1 was detected for chlorophyll fluorescence parameters that explained up to 19.6 % of phenotypic variation. The common QTLs located on chromosomes 7A.1, 4R and 5R, clearly indicated physiological and genetic relationship of the plant survival after freezing with the ability to maintain optimal photochemical activity of the photosystem II and preservation of the cell membranes integrity. The genes located in silico in the identified QTLs include those encoding transmembrane helix proteins like potassium channel and phosphoric ester hydrolase involved in response to osmotic stress as well as proteins involved in the regulation of the gene expression, chloroplast RNA processing and pyrimidine salvage pathway. Additionally, our results confirm that the JIP test is a valuable tool to evaluate freezing tolerance of triticale under unstable winter environments.