QTL mapping for downy mildew resistance in cucumber via bulked segregant analysis using next-generation sequencing and conventional methods

Key message

QTL mapping using NGS-assisted BSA was successfully applied to an F ₂ population for downy mildew resistance in cucumber. QTLs detected by NGS-assisted BSA were confirmed by conventional QTL analysis.

Abstract

Downy mildew (DM), caused by Pseudoperonospora cubensis, is one of the most destructive foliar diseases in cucumber. QTL mapping is a fundamental approach for understanding the genetic inheritance of DM resistance in cucumber. Recently, many studies have reported that a combination of bulked segregant analysis (BSA) and next-generation sequencing (NGS) can be a rapid and cost-effective way of mapping QTLs. In this study, we applied NGS-assisted BSA to QTL mapping of DM resistance in cucumber and confirmed the results by conventional QTL analysis. By sequencing two DNA pools each consisting of ten individuals showing high resistance and susceptibility to DM from a F₂ population, we identified single nucleotide polymorphisms (SNPs) between the two pools. We employed a statistical method for QTL mapping based on these SNPs. Five QTLs, dm2.2, dm4.1, dm5.1, dm5.2, and dm6.1, were detected and dm2.2 showed the largest effect on DM resistance. Conventional QTL analysis using the F₂ confirmed dm2.2 (R ² = 10.8–24 %) and dm5.2 (R ² = 14–27.2 %) as major QTLs and dm4.1 (R ² = 8 %) as two minor QTLs, but could not detect dm5.1 and dm6.1. A new QTL on chromosome 2, dm2.1 (R ² = 28.2 %) was detected by the conventional QTL method using an F₃ population. This study demonstrated the effectiveness of NGS-assisted BSA for mapping QTLs conferring DM resistance in cucumber and revealed the unique genetic inheritance of DM resistance in this population through two distinct major QTLs on chromosome 2 that mainly harbor DM resistance.

     

Induced Bivalent Interlocking and the Course of Meiotic Chromosome Synapsis

WHEN chromosomes pair at meiosis the bivalents so formed do not normally interlock. Heat-treatments can, however, induce bivalent interlocking in the locust Locusta migratoria. Only the longest bivalents interlock and usually only two are found per cell; two “rod” bivalents, with single chiasmata, two “ring” bivalents, each with two or three chiasmata, or one “rod” and one “ring” bivalent (Fig. 1a, b and c). The nature of this interlocking and the metaphase orientational and congressional properties of interlocked bivalents are analysed in detail elsewhere¹.     

Association of three single nucleotide polymorphisms at the SIX1-SIX6 locus with primary open angle glaucoma in the Chinese population

This study investigated the genetic association of three single nucleotide polymorphisms(SNPs; rs10483727, rs33912345, and rs146737847) at the SIX1-SIX6 locus with primary open angle glaucoma(POAG) in the Chinese population. A total of 866 subjects with POAG(685 high-tension glaucoma(HTG) and 181 normal-tension glaucoma(NTG)) and 266 control individuals were included. Significant genetic association was identified for rs10483727 in HTG(P=0.02; odds ratio(OR)=1.31), NTG(P=7.41×10~(-6); OR=2.71), and POAG(i.e., HTG and NTG combined; P=0.001; OR=1.44). rs33912345 was also significantly associated with HTG(P=0.008; OR=1.36), NTG(P=2.72×10~(-6); OR=2.27), and POAG(P=3.84×10~(-4); OR=1.49). The rare SIX6 mutation, rs146737847, was not found in the subjects enrolled in this study. Stratification by patient age identified that both rs10483727 and rs33912345 were significantly associated with NTG in patients aged above 40 years(P=2.08×10~(-5); OR=2.28), whereas in patients aged between 20–40 years, rs33912345 was significantly associated with NTG(P=0.017; OR=2.06). In HTG, the genetic associations for both rs10483727 and rs33912345 were significant in patients aged between 20–40 years(P=0.006; OR=1.56) but not in those aged above 40 years(P=0.118, OR=1.21 and P=0.042, OR=1.29, respectively). This study replicated the association of POAG with two SNPs at the SIX1-SIX6 locus and demonstrated that SNPs, rs10483727 and rs33912345, are significantly associated with POAG, especially with NTG in patients aged above 40 years.     

Congruence and the Biomonitoring of Aquatic Ecosystems: Are Odonate Larvae or Adults the Most Effective for the Evaluation of Impacts

Odonata have been widely used as indicators for the biomonitoring of terrestrial and aquatic habitats due to their sensitivity to environmental impacts. We aimed to determine whether the larval or adult phases of these insects were the best predictors of variation in habitat parameters and the loss of environmental integrity. Specimens were collected during three seasons (dry, rainy, and ebb) from 12 points in the Suiá-missu River basin, at the headwaters of the Xingu River in Mato Grosso, Brazil. The Protest analysis indicated a high degree of congruence between the assemblages of larvae and adults in streams with varying degrees of habitat integrity (R = 0.832, p < 0.001, m ² = 0.307). When the congruence with environmental factors was analyzed, a significant association was found only for the larval phase (R = 0.318, p = 0.03, m ² = 0.888). When the suborders were analyzed separately, congruence was confirmed for anisopteran adults (R = 0.338, p = 0.031, m ² = 0.885) and larvae (R = 0.417, p = 0.003, m ² = 0.826) and for the zygopteran adults (R = 0.345, p = 0.027, m ² = 0.881) and larvae (R = 0.405, p = 0.011, m ² = 0.836). These results indicate that both larvae and adults respond systematically to environmental impacts. We suggest that either life phase can be used for biomonitoring, given their effectiveness for the interpretation of disturbance in terrestrial and aquatic habitats. These findings further reinforce the effectiveness of this insect order for the detection of modifications to the environment, showing that they are good indicators of environmental conditions.     

Simultaneous estimation of saponin contents from soybean seed using Fourier transform infrared spectroscopy and high-performance liquid chromatography analysis

The aim of this study is to determine the feasibility of Fourier transform infrared (FT-IR) spectroscopy for simultaneous determination of saponin contents in different soybean cultivars. In cross validation between predicted content of saponin by PLS regression modeling from FT-IR spectra and measured content by HPLC, total saponin contents were predicted with good accuracy (R ² ≥ 0.71). In external validation, saponin group Ab (R ² = 0.88), saponin DDMP-group βg (R ² = 0.85), total saponin group B (R ² = 0.88), and total saponin content (R ² = 0.87) were predicted with good accuracy, while prediction for saponin group Aa (R ² = 0.58), saponin group Bb′ (R ² = 0.58), and total saponin group A (R ² = 0.25) had relatively lower accuracy. Considering these results, we suggest that the PLS prediction system for saponin contents using FT-IR analysis could be applied as a novel screening tool for high yielding lines in soybean breeding.     

Feed-backs between genetic structure and perturbation-driven decline in seagrass (<Emphasis Type="Italic">Posidonia oceanica</Emphasis>) meadows

We explored the relationships between perturbation-driven population decline and genetic/genotypic structure in the clonal seagrass Posidonia oceanica, subject to intensive meadow regression around four Mediterranean fish-farms, using seven specific microsatellites. Two meadows were randomly sampled (40 shoots) within 1,600 m² at each site: the “impacted” station, 5–200 m from fish cages, and the “control” station, around 1,000 m downstream further away (considered a proxy of the pre-impact genetic structure at the site). Clonal richness (R), Simpson genotypic diversity (D*) and clonal sub-range (CR) were highly variable among sites. Nevertheless, the maximum distance at which clonal dispersal was detected, indicated by CR, was higher at impacted stations than at the respective control station (paired t-test: P < 0.05, N = 4). The mean number of alleles (Â) and the presence of rare alleles ( _r) decreased at impacted stations (paired t-test: P < 0.05, and P < 0.02, respectively, N = 4). At a given perturbation level (quantified by the organic and nutrient loads), shoot mortality at the impacted stations significantly decreased with CR at control stations (R ² = 0.86, P < 0.05). Seagrass mortality also increased with  (R ² = 0.81, P < 0.10), R (R ² = 0.96, P < 0.05) and D* (R ² = 0.99, P < 0.01) at the control stations, probably because of the negative correlation between those parameters and CR. Therefore, the effects of clonal size structure on meadow resistance could play an important role on meadow survival. Large genotypes of P. oceanica meadows thus seem to resist better to fish farm-derived impacts than little ones. Clonal integration, foraging advantage or other size-related fitness traits could account for this effect.     

Inhalable Ipratropium Bromide Particle Engineering with Multicriteria Optimization

Spray-dried ipratropium bromide (IPB) microspheres for oral inhalation were engineered using Quality by Design. The interrogation of material properties, process parameters, and critical product quality attributes interplay enabled rational product design. A 2^7–3 screening design exhibited the Maillard reaction between L-leucine (LL) and lactose at studied outlet temperatures (OT) >130°C. A response surface custom design was used in conjunction with multicriteria optimization to determine the operating design space to achieve inhalable microparticles. Statistically significant predictive models were developed for volume median diameter (p?=?0.0001, adjusted R ²? =?0.9938), span (p?=?0.0278, adjusted R ²? =?0.7912), yield (p?=?0.0020, adjusted R ²? =?0.9320), and OT (p?=?0.0082, adjusted R ²? =?0.8768). An independent verification batch confirmed the model’s predictive capability. The prediction and actual values were in good agreement. Particle size and span were 3.32?±?0.09 μm and 1.71?±?0.18, which were 4.7 and 5.3% higher than the predicted values. The process yield was 50.3%, compared to the predicted value of 65.3%. The OT was 100°C versus the predicted value of 105°C. The label strength of IPB microparticles was 99.0 to 105.9% w/w suggesting that enrichment occurred during the spray-drying process. The present study can be utilized to initiate the design of the first commercial IPB dry powder inhaler.     

Study of the oxidation of alkanes in their mixtures with oxygen and air under the action of a pulsed volume nanosecond discharge

The slow oxidation of alkanes (from methane to hexane) in their stoichiometric mixtures with oxygen or air under the action of nanosecond pulsed discharges was investigated. The discharges were excited in a tube of diameter 5 cm and length of 20 cm by 25-ns voltage pulses with an amplitude of 10 kV and a repetition rate of 40 Hz. The initial pressure in the mixture was varied in the range 0.76–10.1 torr. The current, the electric field strength, and the power deposited in a discharge were measured with a nanosecond time resolution. In time-resolved and time-integrated measurements, the intensities of the following bands were determined: CO ₂ ⁺ (B²Σ → X²Π, δv=0), CH(A²Δ, v′=0 → X²Π, v″=0), OH(A²Σ, v′=0 → X²Π, v″=0), CO(B¹Σ, v′=0 → A¹Π, v″=2), NO(A²Σ → X²Π, δv=3), N₂(C³Π, v′=1 → B³Π, v″=7), N₂(B³Π, v′=6 → A³Σ, v″=3), and N ₂ ⁺ (B²Σ, v′=0 → X²Σ, v″=2). The methane concentration was measured from the absorption of He-Ne laser radiation. Based on the results of optical measurements, the times of the complete oxidation of hydrocarbons were determined.     

<Emphasis Type="SmallCaps">d</Emphasis>-Xylose fermentation,xylitol production and xylanase activities by seven new species of <Emphasis Type="Italic">Sugiyamaella</Emphasis>

Sixteen yeast isolates identified as belonging to the genus Sugiyamaella were studied in relation to D-xylose fermentation, xylitol production, and xylanase activities. The yeasts were recovered from rotting wood and sugarcane bagasse samples in different Brazilian regions. Sequence analyses of the internal transcribed spacer (ITS) region and the D1/D2 domains of large subunit rRNA gene showed that these isolates belong to seven new species. The species are described here as Sugiyamaella ayubii f.a., sp. nov. (UFMG-CM-Y607^T = CBS 14108^T), Sugiyamaella bahiana f.a., sp. nov. (UFMG-CM-Y304^T = CBS 13474^T), Sugiyamaella bonitensis f.a., sp. nov. (UFMG-CM-Y608^T = CBS 14270^T), Sugiyamaella carassensis f.a., sp. nov. (UFMG-CM-Y606^T = CBS 14107^T), Sugiyamaella ligni f.a., sp. nov. (UFMG-CM-Y295^T = CBS 13482^T), Sugiyamaella valenteae f.a., sp. nov. (UFMG-CM-Y609^T = CBS 14109^T) and Sugiyamaella xylolytica f.a., sp. nov. (UFMG-CM-Y348^T = CBS 13493^T). Strains of the described species S. boreocaroliniensis, S. lignohabitans, S. novakii and S. xylanicola, isolated from rotting wood of Brazilian ecosystems, were also compared for traits relevant to xylose metabolism. S. valenteae sp. nov., S. xylolytica sp. nov., S. bahiana sp. nov., S. bonitensis sp. nov., S. boreocarolinensis, S. lignohabitans and S. xylanicola were able to ferment d-xylose to ethanol. Xylitol production was observed for all Sugiyamaella species studied, except for S. ayubii sp. nov. All species studied showed xylanolytic activity, with S. xylanicola, S. lignohabitans and S. valenteae sp. nov. having the highest values. Our results suggest these Sugiyamaella species have good potential for biotechnological applications.     

Meiotic chromosome configurations in oocytes of <Emphasis Type="Italic">Cobitis taenia</Emphasis> and its polyploid hybrids

Diploid and triploid hybrid females of Cobitis as a rule produce unreduced eggs which mainly develop gynogenetically, but some of the eggs incorporate sperm genome and develop into triploids and tetraploids, respectively. Here, we observed for the first time the meiotic chromosomes in the germinal vesicles (GVs) of mature oocytes of three diploid C. taenia (2n = 48) and 20 allopolyploid females of Cobitis (18 triploid 3n = 74 and 2 tetraploid 4n = 99). The majority of GVs in diploid, triploid and tetraploid females contained 24, 74 and 96 or 99 bivalents, respectively. These results directly indicated premeiotic endomitosis as a mechanism underlying the formation of unreduced eggs in allopolyploid females of Cobitis.     

Parasitism Rate of <Emphasis Type="Italic">Habrobracon hebetor</Emphasis> to <Emphasis Type="Italic">Ephestia kuehniella</Emphasis> Larvae: Residual Effect of Deltamethrin,Fenvalerate and Azadirachtin

Sublethal concentrations of chemical insecticides may cause changes in some behavioral characteristics of natural enemies such as functional responses. The residual effect of three synthetic insecticides including deltamethrin, fenvalerate and azadirachtin were studied on functional response of Habrobracon hebetor Say to Ephestia kuehniella Zeller larvae. Seven host densities (2, 4, 8, 16, 32, 64 and 96) were used during a 24 h period. The resulting data were appropriately fit to Type II functional response models in all treatments: (1) control (0.0916 h^?1; and T _h  = 0.2011 h); (2) deltamethrin (a = 0.0839 h^?1; and T _h  = 0.3560 h); (3) fenvalerate (a = 0.0808 h^?1 and T _h  = 0.3623 h); and (4) azadirachtin (a = 0.0900 h^?1 and T _h  = 0.2042 h). Maximum theoretical parasitism rate (T/T _h ) was 119.34 estimated for control wasps. There was no significant difference between the values of attack rates (a and a + D _a ) in all treatments while the handling time was statistically affected in female wasps treated with fenvalerate. Our findings will be useful in safe application of these insecticides in pest management programmes.     

Occurrence and distribution of multiple antibiotic-resistant <Emphasis Type="Italic">Enterococcus</Emphasis> and <Emphasis Type="Italic">Lactobacillus</Emphasis> spp. from Indian poultry: in vivo transferability of their erythromycin,tetracycline and vancomycin resistance

The objective of this study was to determine the occurrence and distribution of antibiotic resistant (AR) lactic acid bacteria (LAB) in Indian poultry. LAB from poultry farm feces (n = 21) and samples from slaughter houses comprising chicken intestine (n = 46), raw meat (n = 23), and sanitary water (n = 4) were evaluated and compared with those from organic chicken (OC) collected from nearby villages. Screening studies showed 5–7 log units higher erythromycin (ER), tetracycline (TC) and vancomycin (VAN) resistant LAB from conventional poultry chicken (CC) compared to OC. Molecular characterization of isolated cultures (n = 32) with repetitive-PCR profiling and 16S rRNA gene sequencing revealed their taxonomical status as Enterococcus faecium (n = 16), Enterococcus durans (n = 2), Lactobacillus plantarum (n = 10), Lactobacillus pentosus (n = 1) and Lactobacillus salivarius (n = 3). The isolates were found to harbor erm(B), msr(C), msr(A/B), tet(M), tet(L) and tet(K) genes associated with Tn916 and Tn917 family transposons. Expression studies through real-time PCR revealed antibiotic-induced expression of the identified AR genes. In vitro and in vivo conjugational studies revealed transfer of ER and TC resistant (ER^R and TC^R) genes with transfer frequencies of 10^?7 and 10^?4 transconjugants recipient^?1, respectively. Although no known VAN resistance (VAN^R) genes were detected, high phenotypic resistance was observed and was transferable to the recipient. From a public health point of view, this study reports Indian poultry as a major source of high levels of AR bacteria contaminating the food chain and the environment. Thus, urgent and determined strategies are needed to control the spread of multiple AR bacteria.     

Molecular cytogenetic characterization of <Emphasis Type="Italic">Triticum timopheevii</Emphasis> chromosomes provides new insight on genome evolution of <Emphasis Type="Italic">T. zhukovskyi</Emphasis>

Triticum timopheevii (2n = 4x = 28, GGA^tA^t) is a tetraploid wheat formerly cultivated in western Georgia. The natural allopolyploid Triticum zhukovskyi is a hexaploid taxon originated from hybridization of T. timopheevii with cultivated einkorn T. monococcum (2n = 2x = 14, A^mA^m). Karyotypically T. timopheevii and T. zhukovskyi differ from other tetraploid and hexaploid wheats and were assigned to the section Timopheevii of the genus Triticum L. Triticum timopheevii and T. zhukovskyi are resistant to many fungal diseases and therefore could potentially be utilized for wheat improvement. We were aiming to precisely identify all T. timopheevii chromosomes and to trace the evolution of T. zhukovskyi. For this, we developed a set of molecular cytogenetic landmarks based on eleven DNA probes. Each chromosome can now be characterized by two to eight probes. The pTa-535 sequence allows the identification of all A^t-genome chromosomes, whereas G-genome and some A^t-genome chromosomes can be identified using (GAA/CTT) _n and pSc119.2 probes. The probes pAesp_SAT86, pAs1, Spelt-1, Spelt-52 and 5S and 45S rDNA can be applied as additional markers to discriminate particular chromosomes or chromosomal regions. The distribution of (GAA/CTT) _n , pTa-535 and pSc119.2 DNA probes on T. timopheevii chromosomes is distinct from other tetraploid wheats and can therefore be used to track individual chromosomes in introgression programs. Our study confirms the origin of T. zhukovskyi from hybridization of T. timopheevii with T. monococcum; however, we show that the emergence was accompanied by changes involving mostly A^t-genome chromosomes. This may be due to the presence of two closely related A-genomes in the T. zhukovskyi karyotype.     

Comparative cytomolecular analyses reveal karyotype variability related to biogeographic and species richness patterns in Bombacoideae (Malvaceae)

Bombacoideae is one out of nine subfamilies of Malvaceae and encompasses 160 tree species. The subfamily is karyotypically characterized by small and numerous chromosomes and is traditionally known by a remarkable inter- and intraspecific chromosome number variation. We conducted a comparative cytogenetic analysis to investigate karyotype diversity and chromosome evolution within Bombacoideae. To achieve this, we performed new chromosome counts, CMA/DAPI double staining, genome size estimations, and localization of 5S and 45S rDNA by fluorescence in situ hybridization for 21 species distributed across the Bombacoideae phylogeny. We performed ancestral states reconstruction analyses to elucidate chromosome evolution and provide insights into the systematics and evolution of Bombacoideae in comparison with other Malvaceae species. Newly generated data on chromosome number on Bombacoideae revealed diploids (Ochroma (2n = 84), Cavanillesia, Pochota, Pseudobombax (2n = 88), and Pachira (2n = 92)) and polyploids (Adansonia digitata (2n = 160) and Eriotheca species (2n = ca. 194 and 2n = 276)). For most species, in situ hybridization revealed karyotype, with two pairs of 45S rDNA sites co-located with CMA⁺ bands, and 5S rDNA sites in only one chromosome pair. Taken together, our results provide support to the hypothesis of karyotypic stability in Bombacoideae. Only the Pachira s.l. clade displayed some variability in ploidy level, number of CMA⁺ bands and 45S rDNA sites, and genome size compared to other Bombacoideae clades. The Striated bark clade was characterized by comparatively small genomes and low cytomolecular variability. Karyotypic data were related to biogeographic and species richness patterns of Bombacoideae.     

Clinical and Microbiological Investigation of Fungemia from Four Hospitals in China

In this study, fungemia cases from four tertiary hospitals located in Shanghai and Anhui province in China from March 2012 to December 2013 were enrolled to investigate clinical features, species distribution, antifungal susceptibility and strain relatedness. During the study period, 137 non-duplicate cases and their corresponding isolates were collected. Six different genera of fungi were identified, of which Candida spp. were the most common (126/137, 91.97 %), with C. albicans predominating (48/137, 35.03 %). The non-Candida fungi rate reached 8.03 % (11/137), and Pichia spp. was the most common (5/137, 3.65 %). Compared with C. albicans, non-C. albicans fungi-associated fungemia was more likely in younger (p = 0.004) and male patients (χ ² = 6.2618, p = 0.0123) and patients from ICUs (χ ² = 6.3783, p = 0.0116). Overall, the susceptible/WT rates of common Candida spp. to fluconazole, itraconazole, voriconazole, flucytosine, amphotericin B and caspofungin were 74.63, 92.31, 93.16, 96.58, 100 and 95.69 %, respectively. C. tropicalis and C. guilliermondii had a low susceptibility to fluconazole: 79.95 and 77.78 %, respectively. No isolates were resistant/WT to caspofungin, but C. parapsilosis and C. guilliermondii had high MIC₉₀ values; 1 and 2 mg/L, respectively. In terms of genotyping, MLST was taken for C. glabrata and C. tropicalis, while microsatellite marker analysis was used for C. albicans and C. parapsilosis. C. glabrata was predominantly clone ST7, accounting for 75 %, while the other isolates showed genetic diversity. Considering the increased proportion of non-C. albicans fungi and the presence of endemic clones of C. glabrata, it is essential to undertake additional surveillance of fungemia.     

Coinheritance of Chromosomes 3 and 11 in the Patients with Autosomal Recessive Polycythemia from Chuvachia

Inheritance of chromosomes 3 and 11 in the families with Chuvash autosomal recessive polycythemia and in control group with no disease symptoms was examined using polymorphic dinucleotide markers D3S1597 and D3S1263, mapped to region 3p25, and D11S4111, D11S4127, and D11S1356, mapped to region 11q23. All patients were homozygous for the C598T mutation in the VHL gene (3p25). The analysis showed that in 75% of the cases, chromosome 3 carrying C598T mutation was coinherited with certain chromosome 11, which differed from 50%, expected upon independent inheritance of each chromosome. In case of chromosome 3 without C598T mutation, this pattern was observed neither in healthy sibs form the families with autosomal recessive polycythemia (44%), nor in the control group (43%). These results suggest that in case of the C598T mutation in the VHL gene, chromosomal loci 3p25 and 11q23 are inherited not independently, compared to the inheritance of these loci in the absence of the mutation in healthy sibs from the affected families χ² = 16.14, p < 0.001), and also in the control family sample (χ² = 17.91, p < 0.001).     

Genetic Polymorphism of <Emphasis Type="Italic">GABRR2</Emphasis> Modulates Individuals’ General Cognitive Ability in Healthy Chinese Han People

Previous studies have indicated that the cognitive impairment or deficit is associated with GABAergic signaling in central nervous system. Inspired by the finding that receptor GABRR2 modulates concentration of GABA and phasic inhibitory GABAergic transmission in brain. This study investigated to what extent a genetic variant (c.1423C>T, rs282129) of GABRR2 gene modulates individuals’ general cognitive ability in 987 Chinese Han people. Results showed a significant influence of GABRR2 gene polymorphism on individuals’ Raven’s Standard Progressive Matrices (RSPM) performance (F = 3.58, P = .028 by ANOVA and χ ² = 9.35, P = .009 by K–W test, respectively), even if non-genetic factors were partialed out (gender, major, types of birthplace, and socioeconomic index) (B = ?.67, SE = .26, t = 2.63, P = .009). The finding provided a strong evidence, to our knowledge, for the view that genetic variant of GABRR2 gene may contribute to the difference of individuals’ general cognitive ability, independently.     

Oxygen transfer as a tool for fine-tuning recombinant protein production by <Emphasis Type="Italic">Pichia pastoris</Emphasis> under glyceraldehyde-3-phosphate dehydrogenase promoter

Effects of oxygen transfer on recombinant protein production by Pichia pastoris under glyceraldehyde-3-phosphate dehydrogenase promoter were investigated. Recombinant glucose isomerase was chosen as the model protein. Two groups of oxygen transfer strategies were applied, one of which was based on constant oxygen transfer rate where aeration rate was Q _O/V = 3 and 10 vvm, and agitation rate was N = 900 min^?1; while the other one was based on constant dissolved oxygen concentrations, C _DO = 5, 10, 15, 20 and 40 % in the fermentation broth, by using predetermined exponential glucose feeding with μ _o = 0.15 h^?1. The highest cell concentration was obtained as 44 g L^?1 at t = 9 h of the glucose fed-batch phase at C _DO = 20 % operation while the highest volumetric and specific enzyme activities were obtained as 4440 U L^?1 and 126 U g^?1 cell, respectively at C _DO = 15 % operation. Investigation of specific enzyme activities revealed that keeping C _DO at 15 % was more advantageous with an expense of relatively higher by-product formation and lower specific cell growth rate. For this strategy, the highest oxygen transfer coefficient and oxygen uptake rate were K _L a = 0.045 s^?1 and OUR = 8.91 mmol m^?3 s^?1, respectively.     

Electronic fine structure calculation of metal complexes with three-open-shell <Emphasis Type="Italic">s</Emphasis>, <Emphasis Type="Italic">d,and p</Emphasis> configurations

The ligand field density functional theory (LFDFT) algorithm is extended to treat the electronic structure and properties of systems with three-open-shell electron configurations, exemplified in this work by the calculation of the core and semi-core 1s, 2s, and 3s one-electron excitations in compounds containing transition metal ions. The work presents a model to non-empirically resolve the multiplet energy levels arising from the three-open-shell systems of non-equivalent ns, 3d, and 4p electrons and to calculate the oscillator strengths corresponding to the electric-dipole 3d ^m  → ns ¹3d ^m 4p ¹ transitions, with n = 1, 2, 3 and m = 0, 1, 2, …, 10 involved in the s electron excitation process. Using the concept of ligand field, the Slater-Condon integrals, the spin-orbit coupling constants, and the parameters of the ligand field potential are determined from density functional theory (DFT). Therefore, a theoretical procedure using LFDFT is established illustrating the spectroscopic details at the atomic scale that can be valuable in the analysis and characterization of the electronic spectra obtained from X-ray absorption fine structure or electron energy loss spectroscopies.     

Characterisation of <Emphasis Type="Italic">Thinopyrum bessarabicum</Emphasis> chromosomes through genome-wide introgressions into wheat

Key message

Genome-wide introgressions of Thinopyrum bessarabicum into wheat resulted in 12 recombinant lines. Cytological and molecular techniques allowed mapping of 1150 SNP markers across all seven chromosomes of the J genome.

Abstract

Thinopyrum bessarabicum (2n = 2x = 14, JJ) is an important source for new genetic variation for wheat improvement due to its salinity tolerance and disease resistance. Its practical utilisation in wheat improvement can be facilitated through development of genome-wide introgressions leading to a variety of different wheat–Th . bessarabicum translocation lines. In this study, we report the generation of 12 such wheat–Th . bessarabicum recombinant lines, through two different crossing strategies, which were characterized using sequential single colour and multi-colour genomic in situ hybridization (sc-GISH and mc-GISH), multi-colour fluorescent in situ hybridization (mc-FISH) and single nucleotide polymorphic (SNP) DNA markers. We also detected 13 lines containing different Th. bessarabicum chromosome aberrations through sc-GISH. Through a combination of molecular and cytological analysis of all the 25 lines containing Th. bessarabicum recombinants and chromosome aberrations we were able to physically map 1150 SNP markers onto seven Th. bessarabicum J chromosomes which were divided into 36 segmental blocks. Comparative analysis of the physical map of Th. bessarabicum and the wheat genome showed that synteny between the two species is highly conserved at the macro-level and confirmed that Th. bessarabicum contains the 4/5 translocation also present in the A genome of wheat. These wheat–Th . bessarabicum recombinant lines and SNP markers provide a useful genetic resource for wheat improvement with the latter having a wider impact as a tool for detection of introgressions from other Thinopyrum species containing the J or a closely-related genome such as Thinopyrum intermedium (J^rJ^rJ^vsJ^vsStSt) and Thinopyrum elongatum (E^eE^e), respectively.