Habtamu Ayalew

Development of a simple and effective silver staining protocol for detection of DNA fragments

Silver staining is one of the widely used methods for DNA fragment detection in biological research. Silver staining protocols have been steadily optimized to improve detection efficiency. This research reports a continuous effort to simplify the existing silver staining protocols, lower experiment cost, and improve DNA detection sensitivity and image clarity. The new method only requires three reagents (silver nitrate, sodium hydroxide, and formaldehyde) and 6–7 min with high detection sensitivity to visualize as low as 14.6 pg (3.3 pg/mm2) of DNA in a non‐denaturing polyacrylamide gel. In comparison to previous reported protocols, the new one has the highest resolution, is the easiest to operate, takes the shortest time, and uses the fewest chemical reagents. Therefore, the new method can be used for quick generation of high quality molecular marker data in genetic analysis.

Quantitative analysis of gene actions controlling root length under water stress in spring wheat (Triticum aestivum L.) genotypes

Abstract. Understanding the genetic control of agronomic traits is important in designing crop improvement programs. Study was conducted to analyse the genetic control of root length under water stress. A full diallel cross of four spring wheat lines, along with their F1 progenies was evaluated under –0.82 MPa water stress in a hydroponic culture. Analysis of variance showed highly significant (P < 0.01) difference among the parental lines and their F1 progenies. Genotypes Santa Elena, Colotana 296–52 and Pato showed comparable longer roots whereas Tincurrin grew significantly (P < 0.05) shorter roots. Genotypes with long roots were found to have more dominant genes than those with shorter roots. Both general and specific combining abilities were highly significant (P < 0.01) indicating the importance of additive and dominant gene effects in the control of root length under water stress. Genotype Santa Elena was found to be the best general combiner whereas the specific cross Santa Elena × Pato was the best hybrid. Moderate narrow-sense heritability (38%) was observed indicating the possibility of improving root length under water stress. The highly significant specific combining ability value (dominant genetic control) suggests that genotypes with more dominant genes should be selected as parents for hybridisation and the hybrid wheat approach might be helpful in improving water stress resistance.

Cell membrane stability and chlorophyll content variation in wheat (Triticum aestivum) genotypes under conditions of heat and drought

Abstract. Heat and drought are among the major obstacles confronting crop production under climate change. The present study was conducted to evaluate 50 diverse wheat genotypes for cell membrane stability (CMS) and chlorophyll content at seedling and anthesis stages under heat and drought stress conditions, to understand the effect of the two abiotic factors and to find promising genotypes for future breeding. Experiments were conducted in the glasshouse (seedling stage) and the field (anthesis stage). Analysis of variance showed significant variation (P ≤ 0.05) for all of the traits at seedling and anthesis stages. High levels of broad-sense heritability and genetic advance at 5% selection intensity indicated the presence of a high genetic component of variation and potential for genetic improvement through selection among the existing genetic variation. CMS showed a significant positive correlation with 1000-grain weight (TGW) under heat and drought conditions at both seedling and anthesis stages. Chlorophyll a/b ratio at seedling stage exhibited a significant negative correlation (r = –0.39, P < 0.05) with TGW under heat stress. Total chlorophyll content was significantly (r = 0.42, P < 0.05) correlated with TGW under heat stress at anthesis. Genotypes ETAD248 and ETAD7 showed the highest CMS and TGW values, whereas their chlorophyll a/b values were lowest, at both seedling and anthesis stages under heat and drought stress conditions. Higher CMS and total chlorophyll content, and lower chlorophyll a/b, were found to be useful indicators to identify genotypes with high TGW under heat and drought stress conditions. This study indicated the possibility of using seedling resistance as an indicator for later stage response in breeding for heat and drought resistance. The resistant genotypes identified can be used as potential germplasm in breeding programs.

Identification of Early Vigor QTLs and QTL by Environment Interactions in Wheat ( Triticum eastivum L.)

Crop productivity is highly dependent on successful seed germination and seedling establishment. This study evaluated two mapping populations, Batavia/Ernie (double haploid) and Synthetic/Opata (recombinant inbred lines), for early vigor under water stress and normal growing conditions. Significant gene, environment (water), and gene by environment interaction effects were observed. Broad sense heritability was 29 and 40% for the Batavia/Ernie and Synthetic/Opata populations, respectively. Quantitative trait loci (QTL) were analyzed based on single and multienvironment models. The two mapping populations differed in the number and locations of QTLs except qNev.uwa.4AL was identified in both populations under the non-stress condition, while qSev.uwa.3BL was specifically expressed under the stress condition in the Synthetic/Opata population. QTL by environment interaction (QEI) enabled identification of nine QTLs, including those identified by the single environment approach. Phenotypic variation expression (PVE) of QEIs ranged from 4.8 to 14.9% across the populations. Larger proportion of PVE of QEIs was explained by the additive components. Favorable alleles for three of the QTLs identified in the Synthetic/Opata population were derived from Synthetic, while Batavia contributed favorable alleles to a QTL on the long arm of chromosome 1D in the Batavia/Ernie population. QTL detected under water stress (qSev.uwa.3BL) co-located with the DREB 1 gene, which was mapped between markers Xmwg818 and Xfbb117 (58.1–77.6 cM). QTLs with high proportion of additive components can be validated for marker assisted gene pyramiding and selection.

Identification and validation of a major chromosome region for high grain number per spike under meiotic stage water stress in wheat (Triticum aestivum L.)

Grain number is a major trait for wheat yield under dryland farming. An International Triticeae Mapping Initiative (ITMI) mapping population comprising 105 recombinant inbred lines (RIL) developed from a cross between a Synthetic hexaploid wheat (Triticum aestivum) ‘W7984’ and a spring wheat variety ‘Opata M85’ was used to identify quantitative trait loci (QTL) associated with grain number per spike under two treatment conditions, normal watering and water stress during meiosis. Two major QTL for grain number per spike on the main stem Q.Gnu.uwa-5A-1 and Q.Gnu.uwa-5A-2 with phenotypic variations of 25.71% and 24.93%, respectively, were detected on the long arm of chromosome 5A when plants were exposed to water stress during meiosis. One QTL (Q.Gnu.uwa-2A) with a LOD score of 2.8 was detected on the long arm of chromosome 2A under normal watering condition. The alleles associated with higher grain number per spike under different treatment conditions came from the Synthetic W7984 parent. Two populations developed from crosses Synthetic W7984 × Lang and Synthetic W7984 × Westonia were used to validate the identified QTL under water stress during meiosis. SSR markers Xbarc230 and Xbarc319 linked with the identified QTL on chromosome 5AL were validated in the two F2:4 segregating populations. These closely linked SSR markers could potentially be utilized in marker-assisted selection to reduce yield loss in regions where water stress during meiosis occurs frequently. The identified QTL can be incorporated into elite lines / cultivars to improve wheat grain yield.