Combined detection and introgression of QTL in outbred populations

Background

Detecting a QTL is only the first step in genetic improvement programs. When a QTL with desirable characteristics is found, e.g. in a wild or unimproved population, it may be interesting to introgress the detected QTL into the commercial population. One approach to shorten the time needed for introgression is to combine both QTL identification and introgression, into a single step. This combines the strengths of fine mapping and backcrossing and paves the way for introgression of desirable but unknown QTL into recipient animal and plant lines.

Methods

The method consisting in combining QTL mapping and gene introgression has been extended from inbred to outbred populations in which QTL allele frequencies vary both in recipient and donor lines in different scenarios and for which polygenic effects are included in order to model background genes. The effectiveness of the combined QTL detection and introgression procedure was evaluated by simulation through four backcross generations.

Results

The allele substitution effect is underestimated when the favourable QTL allele is not fixed in the donor line. This underestimation is proportional to the frequency differences of the favourable QTL allele between the lines. In most scenarios, the estimates of the QTL location are unbiased and accurate. The retained donor chromosome segment and linkage drag are similar to expected values from other published studies.

Conclusions

In general, our results show that it is possible to combine QTL detection and introgression even in outbred species. Separating QTL mapping and introgression processes is often thought to be longer and more costly. However, using a combined process saves at least one generation. With respect to the linkage drag and obligatory drag, the results of the combined detection and introgression scheme are very similar to those of traditional introgression schemes.     

Variation in the Essential Oil Composition,Antioxidant Capacity,and Physiological Characteristics of Pelargonium graveolens L. Inoculated with Two Species of Mycorrhizal Fungi Under Water Deficit Conditions

In recent years, the application of arbuscular mycorrhizal fungi (AMF) has been considered to be an important strategy for improving crop yield and quality. In the present study, a factorial experiment based on a complete randomized design with two factors was performed to investigate the effect of AMF and water stress on the essential oil (EO) composition, antioxidant activity, and physiological and morphological characteristics of rose-scented geranium (Pelargonium graveolens L.). The factors included AMF inoculation (Rhizophagus intraradices, Funneliformis mosseae, and a mixture of both species) and irrigation levels [well-watered (WW), moderate drought stress (MDS), and severe drought stress (SDS)]. The main EO constituents were citronellol (31–37%) and geraniol (9–14%) in all treatments. Under water-stress conditions, some constituents increased, such as geraniol and geranyl formate, whereas others decreased, such as linalool, menthone and rose oxide. Overall, the highest amount of citronellol (37.3%) and geraniol (14.8%) was obtained in the plants inoculated with F. mosseae and R. intraradices under WW and MDS conditions, respectively. Antioxidant activity, total flavonoids, and phenolics were increased because of AMF inoculation, whereas a different trend was observed for the phenolic and flavonoid contents under water-stress conditions. Furthermore, water deficit elevated the amount of soluble carbohydrates as well as the proline content, whereas the amount of proline was lower in inoculated plants than in non-inoculated ones. All the growth parameters were improved in the AMF-inoculated plants compared to non-inoculated ones under different irrigation regimes. Drought conditions decreased the photosynthetic pigments and efficiency, whereas AMF plants ameliorated the adverse effect of drought conditions. In general, mycorrhizal inoculation resulted in an improvement in the growth parameters as well as the phytochemical and physiological characteristics of rose-scented geranium.

     

Thermodynamic of Water Sorption of Grape Seed: Temperature Effect of Sorption Isotherms and Thermodynamic Characteristics

After determination of sorption isotherms of grape seeds using gravimetric method, five models with temperature effect were used to fit water sorption isotherms of grape seeds to investigate temperature effect on sorption isotherms and its thermodynamic characteristics. Halsey model had minimum mean relative percentage error (M _e ) and all other models used were good in fitting experimental data (with M _e of less than 10 %). Differential parameters such as net isosteric heat, isosteric heat, differential entropy and integral function such as equilibrium heat, net equilibrium heat, integral entropy and surface potential have been calculated. The net isosteric heat, isosteric heat and differential entropy decreased with moisture content. The net equilibrium enthalpy, equilibrium enthalpy and integral entropy decreased with moisture content. The surface potential at four temperatures (35, 45, 55 and 65 °C) was estimated, and low temperature effect was reported.     

Galactosylated collagen matrix enhanced in vitro maturation of human embryonic stem cell-derived hepatocyte-like cells

Due to their important biomedical applications, functional human embryonic stem cell-derived hepatocyte-like cells (hESC-HLCs) are an attractive topic in the field of stem cell differentiation. Here, we have initially differentiated hESCs into functional hepatic endoderm (HE) and continued the differentiation by replating them onto galactosylated collagen (GC) and collagen matrices. The differentiation of hESC-HE cells into HLCs on GC substrate showed significant up-regulation of hepatic-specific genes such as ALB, HNF4α, CYP3A4, G6P, and ASGR1. There was more albumin secretion and urea synthesis, as well as more cytochrome p450 activity, in differentiated HLCs on GC compared to the collagen-coated substrate. These results suggested that GC substrate has the potential to be used for in vitro maturation of hESC-HLCs.     

Viscoelasticity of Tau Proteins Leads to Strain Rate-Dependent Breaking of Microtubules during Axonal Stretch Injury: Predictions from a Mathematical Model

The unique viscoelastic nature of axons is thought to underlie selective vulnerability to damage during traumatic brain injury. In particular, dynamic loading of axons has been shown to mechanically break microtubules at the time of injury. However, the mechanism of this rate-dependent response has remained elusive. Here, we present a microstructural model of the axonal cytoskeleton to quantitatively elucidate the interaction between microtubules and tau proteins under mechanical loading. Mirroring the axon ultrastructure, the microtubules were arranged in staggered arrays, cross-linked by tau proteins. We found that the viscoelastic behavior specifically of tau proteins leads to mechanical breaking of microtubules at high strain rates, whereas extension of tau allows for reversible sliding of microtubules without any damage at small strain rates. Based on the stiffness and viscosity of tau proteins inferred from single-molecule force spectroscopy studies, we predict the critical strain rate for microtubule breaking to be in the range 22–44 s⁻¹, in excellent agreement with recent experiments on dynamic loading of micropatterned neuronal cultures. We also identified a characteristic length scale for load transfer that depends on microstructural properties and have derived a phase diagram in the parameter space spanned by loading rate and microtubule length that demarcates those regions where axons can be loaded and unloaded reversibly and those where axons are injured due to breaking of the microtubules.     

Incorporation of SPION-casein core-shells into silk-fibroin nanofibers for cardiac tissue engineering

Mimicking the structure of extracellular matrix (ECM) of myocardium is necessary for fabrication of functional cardiac tissue. The superparamagnetic iron oxide nanoparticles (SPIONs, Fe₃O₄), as new generation of magnetic nanoparticles (NPs), are highly intended in biomedical studies. Here, SPION NPs (1 wt%) were synthesized and incorporated into silk-fibroin (SF) electrospun nanofibers to enhance mechanical properties and topography of the scaffolds. Then, the mouse embryonic cardiac cells (ECCs) were seeded on the scaffolds for in vitro studies. The SPION NPs were studied by scanning electron microscope (SEM), X-ray diffraction (XRD), and transmission electron microscope (TEM). SF nanofibers were characterized after incorporation of SPIONs by SEM, TEM, water contact angle measurement, and tensile test. Furthermore, cytocompatibility of scaffolds was confirmed by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. SEM images showed that ECCs attached to the scaffolds with elongated morphologies. Also, the real-time PCR and immunostaining studies approved upregulation of cardiac functional genes in ECCs seeded on the SF/SPION-casein scaffolds including GATA-4, cardiac troponin T, Nkx 2.5, and alpha-myosin heavy chain, compared with the ones in SF. In conclusion, incorporation of core-shells in SF supports cardiac differentiation, while has no negative impact on ECCs' proliferation and self-renewal capacity.     

Production of bifunctional single-chain antibody-based fusion proteins in <Emphasis Type="Italic">Pichia </Emphasis><Emphasis Type="Italic">pastoris</Emphasis> supernatants

Recombinant antibody fusion constructs with heterologous functional domains are a promising approach to new therapeutic targeting strategies. However, expression of such constructs is mostly limited to cost and labor-intensive mammalian expression systems. Here we report on the employment of Pichia pastoris for the expression of heterologous antibody fusion constructs with green fluorescent protein, A33scFv::GFP, or with cytosine deaminase, A33scFv::CDy, their production in a biofermenter and a modified purification strategy. Combined, these approaches improved production yields by about thirty times over established standard protocols, with extracellular secretion of the fusion construct reaching 12.0 mg/l. Bifunctional activity of the fusion proteins was demonstrated by flow cytometry and an in-vitro cytotoxicity assay. With equal amounts of purified protein, the modified purification method lead to higher functional results. Our results demonstrate the suitability of methylotrophic Pichia expression systems and laboratory-scale bioreactors for the production of high quantities of bifunctionally active heterologous single-chain fusion proteins.     

Multiplexed genotyping with sequence-tagged molecular inversion probes

We report on the development of molecular inversion probe (MIP) genotyping, an efficient technology for large-scale single nucleotide polymorphism (SNP) analysis. This technique uses MIPs to produce inverted sequences, which undergo a unimolecular rearrangement and are then amplified by PCR using common primers and analyzed using universal sequence tag DNA microarrays, resulting in highly specific genotyping. With this technology, multiplex analysis of more than 1,000 probes in a single tube can be done using standard laboratory equipment. Genotypes are generated with a high call rate (95%) and high accuracy (>99%) as determined by independent sequencing.     

Natural parasitism of the diamondback moth,Plutella xylostella (L.) (Lep.: Plutellidae) by a larval parasitoid wasp,Diadegma anurum on different cauliflower cultivars

The diamondback moth (DBM), Plutella xylostella (L.) (Lepidoptera: Yponomeutidae), is the most serious pest of cauliflower fields in central Iran and its control is primarily based on pesticide sprays. Over the past years, a number of new pesticide compounds were introduced onto the market and some of them may cause adverse effects on natural populations of parasitoids associated with DBM. Excessive use of insecticides against the pest did not produce satisfactory results but has caused concerns about environmental pollution and increased pest resistance to chemicals. This research aims to study natural parasitism of pest on different cauliflower cultivars in the fields of south of Tehran. Dominant species of parasitoids include Diadegma anurum, Cotesia plutellae and Oomyzus sokolowskii. The highest parasitism rate was observed by D. anurum that was recorded on Buris cultivar (19.92?±?1.06) and White cloud cultivar (16.20?±?1.49) and the lowest parasitism rate was observed on Snow crown cultivar (3.42) and SG cultivar (5.00) during the season.