Simulation Experiments on Efficiencies of Gene Introgression by Backcrossing

Designing a highly efficient backcross (BC) marker-assisted selection (MAS) experiment is not a straightforward exercise, efficiency being defined here as the ratio between the resources that need to be invested at each generation and the number of generations required to achieve the selection. This paper presents results of simulations conducted for different strategies, using the maize genome as a model, to compare allelic introgression with DNA markers through BCs. Simulation results indicate that the selection response in the BC1 could be increased significantly when the selectable population size (N sl) is 100. Selectable population size is defined as the number of individuals with favorable alleles at the target loci from which selection with markers can be carried out on the rest of the genome at nontarget loci, simulations considered the allelic introgression at one to five target loci, with different population sizes, changes in the recombination frequency between target loci and flanking markers, and different numbers of genotypes selected at each generation. For an introgression at one target locus in a partial line conversion, and using MAS at nontarget loci only at one generation, a selection at BC3 would be more efficient than a selection at BC1 or BC2, due to the increase over generations of the ratio of the standard deviation to the mean of the donor genome contribution. With selection only for the presence of a donor allele at one locus in BC1 and BC2, and MAS at BC3, lines with <5% of the donor genome can be obtained with a N sl of 10 in BC1 and BC2, and 100 in BC3 These results are critical in the application of molecular markers to introgress elite alleles as part of plant improvement program

Identification of quantitative trait loci under drought conditions in tropical maize. 2. Yield components and marker-assisted selection strategies

In most maize-growing areas yield reductions due to drought have been observed. Drought at flowering time is, in some cases, the most damaging. In the experiment reported here, trials with F3 families, derived from a segregating F2 population, were conducted in the field under well-watered conditions (WW) and two other water-stress regimes affecting flowering (intermediate stress, IS, and severe stress, SS). Several yield components were measured on equal numbers of plants per family: grain yield (GY), ear number (ENO), kernel number (KNO), and 100-kernel weight (HKWT). Correlation analysis of these traits showed that they were not independent of each other. Drought resulted in a 60% decrease of GY under SS conditions. By comparing yield under WW and SS conditions, the families that performed best under WW conditions were found to be proportionately more affected by stress, and the yield reductions due to SS conditions were inversely proportional to the performance under drought. Moreover, no positive correlation was observed between a drought-tolerance index (DTI) and yield under WW conditions. The correlation between GY under WW and SS conditions was 0.31. Therefore, in this experiment, selection for yield improvement under WW conditions only, would not be very effective for yield improvement under drought. Quantitative trait loci (QTLs) were identified for GY, ENO and KNO using composite interval mapping (CIM). No major QTLs, expressing more then 13% of the phenotypic variance, were detected for any of these traits, and there were inconsistencies in their genomic positions across water regimes. The use of CIM allowed the evaluation of QTL-by-environment interactions (Q×E) and could thus identify “stable” QTLs CIMMYT, Apartado Postal 6-641, 06600 Mexico D.F., Mexico across drought environments. Two such QTLs for GY, on chromosomes 1 and 10, coincided with two stable QTLs for KNO. Moreover, four genomic regions were identified for the expression of both GY and the anthesis-silking interval (ASI). In three of these, the allelic contributions were for short ASI and GY increase, while for that on chromosome 10 the allelic contribution for short ASI corresponded to a yield reduction. From these results, we hypothesize that to improve yield under drought, marker-assisted selection (MAS) using only the QTLs involved in the expression of yield components appears not to be the best strategy, and neither does MAS using only QTLs involved in the expression of ASI. We would therefore favour a MAS strategy that takes into account a combination of the “best QTLs” for different traits. These QTLs should be stable across target environments, represent the largest percentage possible of the phenotypic variance, and, though not involved directly in the expression of yield, should be involved in the expression of traits significantly correlated with yield, such as ASI

role of winter host plants in vineyard colonization and phenology of zygina rhamni hemiptera cicadellidae typhlocybinae

Abstract Zygina rhamni Ferrari (Hemiptera: Cicadellidae: Typhlocybinae) is a mesophyll-feeding leafhopper that infests grapevine, Vitis vinifera L. This leafhopper's overwintering strategy and the dynamics of its vineyard colonization, relative to the distance between the grapevines and the winter host plants and different grapevine vegetative densities, are described herein. This species shows a facultative heteroecious life cycle. In autumn, it migrates from vineyards to bramble, Rubus gr. fruticosus, and rose, Rosa chinensis Jacquin. On these host plants, females lay eggs that start to hatch at the end of March. Adults that developed on bramble colonize grapevines beginning in May; the leaf fall of rose, which takes place during the first half of April, probably causes the death of unhatched eggs and nymphs. We found that overwintering females can move back to vineyards and lay eggs until early June. The dynamics of vineyard colonization by the leafhopper involved initial concentration in areas close t..

Identification of orthologous regions associated with tissue growth under water-limited conditions

Plant recovery from early season drought is related to the amount of biomass retained during stress and biomass production after the end of stress. Reduction in leaf expansion is one of the first responses to water deficit. It is assumed that the control of tissue development under water deficit contributes to traits such as early vigor, as well as maintenance of growth of reproductive organs. To dissect the underlying mechanisms controlling tissue expansion under water-limited conditions, we used a multilevel approach combining quantitative genetics and genomics. To identify orthologous genetic regions controlling tissue growth under water-limited conditions a series of QTL mapping and microarray gene expression studies were conducted in rice and maize. Results of differentially expressed genes from microarray experiments, QTLs and candidate genes related to growth in the different species are compared on consensus maps (within species) and then on synteny maps (between species), to identify common genetic regions between rice and maize

Identification of several small main-effect QTLs and a large number of epistatic QTLs for drought tolerance related traits in groundnut (Arachishypogaea L.)

Cultivated groundnut or peanut (Arachis hypogaea L.), an allotetraploid (2n = 4x = 40), is a self pollinated and widely grown crop in the semi-arid regions of the world. Improvement of drought tolerance is an important area of research for groundnut breeding programmes. Therefore, for the identification of candidate QTLs for drought tolerance, a comprehensive and refined genetic map containing 191 SSR loci based on a single mapping population (TAG 24 × ICGV 86031), segregating for drought and surrogate traits was developed. Genotyping data and phenotyping data collected for more than ten drought related traits in 2–3 seasons were analyzed in detail for identification of main effect QTLs (M-QTLs) and epistatic QTLs (E-QTLs) using QTL Cartographer, QTLNetwork and Genotype Matrix Mapping (GMM) programmes. A total of 105 M-QTLs with 3.48–33.36% phenotypic variation explained (PVE) were identified using QTL Cartographer, while only 65 M-QTLs with 1.3–15.01% PVE were identified using QTLNetwork. A total of 53 M-QTLs were such which were identified using both programmes. On the other hand, GMM identified 186 (8.54–44.72% PVE) and 63 (7.11–21.13% PVE), three and two loci interactions, whereas only 8 E-QTL interactions with 1.7–8.34% PVE were identified through QTLNetwork. Interestingly a number of co-localized QTLs controlling 2–9 traits were also identified. The identification of few major, many minor M-QTLs and QTL × QTL interactions during the present study confirmed the complex and quantitative nature of drought tolerance in groundnut. This study suggests deployment of modern approaches like marker-assisted recurrent selection or genomic selection instead of marker-assisted backcrossing approach for breeding for drought tolerance in groundnut

Phosphoenolpyruvate carboxylase dentified as a key enzyme in erythrocytic Plasmodium falciparum carbon metabolism

Phospoenolpyruvate carboxylase (PEPC) is absent from humans but encoded in thePlasmodium falciparum genome, suggesting that PEPC has a parasite-specific function. To investigate its importance in P. falciparum, we generated a pepc null mutant (D10Δpepc), which was only achievable when malate, a reduction product of oxaloacetate, was added to the growth medium. D10Δpepc had a severe growth defect in vitro, which was partially reversed by addition of malate or fumarate, suggesting that pepc may be essential in vivo. Targeted metabolomics using 13C-U-D-glucose and 13C-bicarbonate showed that the conversion of glycolytically-derived PEP into malate, fumarate, aspartate and citrate was abolished in D10Δpepc and that pentose phosphate pathway metabolites and glycerol 3-phosphate were present at increased levels. In contrast, metabolism of the carbon skeleton of 13C,15N-U-glutamine was similar in both parasite lines, although the flux was lower in D10Δpepc; it also confirmed the operation of a complete forward TCA cycle in the wild type parasite. Overall, these data confirm the CO2 fixing activity of PEPC and suggest that it provides metabolites essential for TCA cycle anaplerosis and the maintenance of cytosolic and mitochondrial redox balance. Moreover, these findings imply that PEPC may be an exploitable target for future drug discovery

Optimized high gradient magnetic separation for isolation of Plasmodium-infected red blood cells

<p>Abstract</p> <p>Background</p> <p>Highly purified infected red blood cells (irbc), or highly synchronized parasite cultures, are regularly required in malaria research. Conventional isolation and synchronization rely on density and osmotic fragility of irbc, respectively. High gradient magnetic separation (HGMS) offers an alternative based on intrinsic magnetic properties of irbc, avoiding exposure to chemicals and osmotic stress. Successful HGMS concentration in malaria research was previously reported using polymer coated columns, while HGMS depletion has not been described yet. This study presents a new approach to both HGMS concentration and depletion in malaria research, rendering polymer coating unnecessary.</p> <p>Methods</p> <p>A dipole magnet generating a strong homogenous field was custom assembled. Polypropylene syringes were fitted with one-way stopcocks and filled with stainless steel wool. Rbc from <it>Plasmodium falciparum </it>cultures were resuspended in density and viscosity optimized HGMS buffers and HGMS processed. Purification and depletion results were analysed by flow cytometer and light microscopy. Viability was evaluated by calculating the infection rate after re-culturing of isolates.</p> <p>Results</p> <p>In HGMS concentration, purity of irbc isolates from asynchronous cultures consistently ranged from 94.8% to 98.4% (mean 95.7%). With further optimization, over 90% of isolated irbc contained segmented schizonts. Processing time was less than 45 min. Reinfection rates ranged from 21.0% to 56.4%. In HGMS depletion, results were comparable to treatment with sorbitol, as demonstrated by essentially identical development of cultures.</p> <p>Conclusion</p> <p>The novel HGMS concentration procedure achieves high purities of segmented stage irbc from standard asynchronous cultures, and is the first HGMS depletion alternative to sorbitol lysis. It represents a simple and highly efficient alternative to conventional irbc concentration and synchronization methods.</p

Promiscuous Expression of α-Tubulin II in Maturing Male and Female Plasmodium falciparum Gametocytes

BACKGROUND: Antimalarial interventions designed to impact on the transmissible sexual stages of Plasmodium falciparum are evaluated by measurement of peripheral gametocyte carriage in vivo and infectivity to mosquitoes. Drug or vaccine-elicited effects may differentially affect the relative abundance of mature male and female sexual forms, and this can be measured by estimation of sex ratios before and after intervention in vivo and in vitro. Measuring the impact of anti-gametocyte drugs on sexual commitment of immature gametocyte stages in vitro is not currently possible as male and female parasites cannot be distinguished by morphology alone prior to stage IV. METHODOLOGY/PRINCIPAL FINDINGS: We have modified an existing immunofluorescence-based approach for distinguishing male and female gametocytes during development in vitro, by using highly synchronised magnetically-enriched gametocyte preparations at different stages of maturity. Antibodies recognising α-tubulin II (males) and Pfg377 (females) were used to attempt to discriminate the sexes. Transcription of these two proteins was not coordinated during in vitro development, with pfg377 transcripts accumulating only late in development, immediately prior to immunofluorescent signals from the PfG377 protein appearing in stage IV gametocytes. Contrary to previous descriptions of this protein as male-specific in P. falciparum, α-tubulin II recognised both male and female gametocytes at stages I to IV, but evidence of differential expression levels of this protein in late stage male and female gametocytes was found. Using antibodies recognising PfG377 as the primary marker and α-tubulin II as a secondary marker, robust estimates of sex ratio in in vitro cultures were obtained for gametocytes at stage IV or later, and validated by light microscopic counts. However, sex ratio estimation was not possible for early stage gametocytes due to the promiscuity of α-tubulin II protein expression, and the relatively late accumulation of PfG377 during the development process. CONCLUSIONS/SIGNIFICANCE: This approach is a feasible method for the evaluation of drug impacts on late-stage gametocyte sex ratio in in vitro studies. Additional sex-specific antigens need to be evaluated for sex ratio estimation in early stage gametocyte preparations

QTL Analysis of Shading Sensitive Related Traits in Maize under Two Shading Treatments

During maize development and reproduction, shading stress is an important abiotic factor influencing grain yield. To elucidate the genetic basis of shading stress in maize, an F2:3 population derived from two inbred lines, Zhong72 and 502, was used to evaluate the performance of six traits under shading treatment and full-light treatment at two locations. The results showed that shading treatment significantly decreased plant height and ear height, reduced stem diameter, delayed day-to-tassel (DTT) and day-to-silk (DTS), and increased anthesis-silking interval (ASI). Forty-three different QTLs were identified for the six measured traits under shading and full light treatment at two locations, including seven QTL for plant height, nine QTL for ear height, six QTL for stem diameter, seven QTL for day-to-tassel, six QTL for day-to-silk, and eight QTL for ASI. Interestingly, three QTLs, qPH4, qEH4a, and qDTT1b were detected under full sunlight and shading treatment at two locations simultaneously, these QTL could be used for selecting elite hybrids with high tolerance to shading and high plant density. And the two QTL, qPH10 and qDTS1a, were only detected under shading treatment at two locations, should be quit for selecting insensitive inbred line in maize breeding procedure by using MAS method