Structural characterization,expression analysis and evolution of the red/far-red sensing photoreceptor gene,phytochrome C (<Emphasis Type="Italic">PHYC</Emphasis>), localized on the ‘B’ genome of hexaploid wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

Phytochromes are a family of red/far-red light perceiving photoreceptors.The monocot phytochrome family is represented by three members, PHYA, PHYB and PHYC. We have isolated and characterized the first PHY gene member (TaPHYC) from common wheat, Triticum aestivum var. CPAN1676. It codes for a species of the photoreceptor, phyC, which is known to be light-stable in all plants analyzed so far. A sequence of 7.2 kb has been determined, which includes 3.42 kb of coding region.This is the second full-length PHYC gene sequenced from a monocot (first was from rice). TaPHYC gene shares structural similarities with the rice PHYC containing four exons and three introns in the coding region. The 5 UTR is 1.0-kb-long and harbors an upstream open reading frame (URF) encoding 28 aa. Southern blot analysis of TaPHYC indicates that it represents single locus in the wheat genome, although the possibility of additional loci cannot be completely ruled out. Chromosomal localization using nullisomic–tetrasomic lines of Triticum aestivum var. Chinese Spring places TaPHYC on chromosome 4B. PHYC represents a constitutively expressed gene in all the organs tested and under light/dark conditions. However, PHYC was found to be developmentally regulated showing maximal expression in 3-day-old dark-grown seedlings, which declined thereafter. In silico analysis has also been done to compare TaPHYC gene with the partial sequences known from other wheat species and cultivars. The presence of a topoisomerase gene immediately downstream of the PHYC gene, both in rice and wheat genomes, presents yet another example of synteny in cereals and its possible significance has been discussed.     

Determining the evolutionary potential of a gene

In addition to information for current functions, the sequence of a gene includes potential information for the evolution of new functions. The wild-type ebgA (evolved beta-galactosidase) gene of Escherichia coli encodes a virtually inactive beta-galactosidase, but that gene has the potential to evolve sufficient activity to replace the lacZ gene for growth on the beta-galactoside sugars lactose and lactulose. Experimental evidence, which has suggested that the evolutionary potential of Ebg enzyme is limited o two specific amino acid replacements, is limited to examining the consequences of single base- substitutions. Thirteen beta-galactosidases homologous with the Ebg beta-galactosidase are widely dispersed, being found in gram-negative and gram-positive eubacteria and in a eukaryote. A comparison of Ebg beta-galactosidase with those 13 beta-galactosidases shows that Ebg is part of an ancient clade that diverged from the paralogous lacZ beta- galactosidase over 2 billion years ago. Ebg differs from other members of its clade at only 2 of the 15 active-site residues, and the two mutations required for full Ebg beta-galactosidase activity bring Ebg into conformity with the other members of its clade. We conclude that either these are the only acceptable amino acids at those positions, or all of the single-base-substitution replacements that must arise as intermediates on the way to other acceptable amino acids are so deleterious that they constitute a deep selective valley that has not been traversed in over 2 billion years. The evolutionary potential of Ebg is thus limited to those two replacements.      

Poor physical fitness is independently associated with mild cognitive impairment in elderly Koreans

The purpose of this study was to investigate the association between physical fitness and mild cognitive impairment (MCI) in elderly Koreans. This was a cross-sectional study that involved 134 men and 299 women aged 65 to 88 years. Six senior fitness tests were used as independent variables: 30 s chair stand for lower body strength, arm curl for upper body strength, chair-sit-and-reach for lower body flexibility, back scratch for upper body flexibility, 8-ft up-and-go for agility/dynamic balance, and 2-min walk for aerobic endurance. Global cognitive function was assessed using the Korean version of the Mini-Mental State Examination (MMSE). Potential covariates such as age, education levels, blood lipids, and insulin resistance (IR) markers were also assessed. Compared to individuals without MMSE-based MCI, individuals with MMSE-based MCI had poor physical fitness based on the senior fitness test (SFT). There were significant positive trends observed for education level (p=0.001) and MMSE score (p<0.001) across incremental levels of physical fitness in this study population. Individuals with moderate (OR=0.341, p=0.006) and high (OR=0.271, p=0.007) physical fitness based on a composite score of the SFT measures were less likely to have MMSE-based MCI than individuals with low physical fitness (referent, OR=1). The strength of the association between moderate (OR=0.377, p=0.038) or high (OR=0.282, p=0.050) physical fitness and MMSE-based MCI was somewhat attenuated but remained statistically significant even after adjustment for the measured compounding factors. We found that poor physical fitness was independently associated with MMSE-based MCI in elderly Koreans.     

Molecular dynamics of synthetic leucine-serine ion channels in a phospholipid membrane

Molecular dynamics calculations were carried out on models of two synthetic leucine-serine ion channels: a tetrameric bundle with sequence (LSLLLSL)(3)NH(2) and a hexameric bundle with sequence (LSSLLSL)(3)NH(2). Each protein bundle is inserted in a palmitoyloleoylphosphatidylcholine bilayer membrane and solvated by simple point charge water molecules inside the pore and at both mouths. Both systems appear to be stable in the absence of an electric field during the 4 ns of molecular dynamics simulation. The water motion in the narrow pore of the four-helix bundle is highly restricted and may provide suitable conditions for proton transfer via a water wire mechanism. In the wider hexameric pore, the water diffuses much more slowly than in bulk but is still mobile. This, along with the dimensions of the pore, supports the observation that this peptide is selective for monovalent cations. Reasonable agreement of predicted conductances with experimentally determined values lends support to the validity of the simulations.     

Identification of eight chromosomes and a microsatellite marker on 1AS associated with QTL for grain weight in bread wheat

The present study in bread wheat was undertaken, firstly, to identify chromosomes carrying QTLs controlling 1000 grain weight (GW) and, secondly, to develop molecular marker(s) linked with this trait. Using the genotype Rye Selection111 (RS111), we carried out a monosomic analysis that suggested that 8 chromosomes (1A, 1D, 2B, 4B, 5B, 6B, 7A and 7D) carried QTLs controlling GW, with only 3 of these (1A, 2B, 7A) carrying alleles for high GW. To tag the QTLs present on these chromosomes, we crossed the genotype RS111 with high GW (56.83 g) with the genotype Chinese Spring (CS) with low GW (23.74 g) and obtained 100 RILs. These RILs showed normal distribution for GW. The parental genotypes were analysed with as many as 346 STMS primer pairs for detection of polymorphism. Of these, 267 primer pairs gave scorable amplification products, 63 of which detected polymorphism between the parents. Using each of these 63 primer pairs, we carried out bulked segregant analysis on RILs representing two extremes of the distribution. One primer pair (WMC333) showed an association of the marker locus Xwmc333 with grain weight. This was confirmed through selective genotyping, and the co-segregation data on molecular marker locus Xwmc333 and GW were analysed following a single marker linear regression approach. Significant regression suggested linkage between Xwmc333 and a QTL for GW. The results showed that the above QTL accounted for 15.09% of the variation for GW between the parents. The marker has been located on chromosome arm 1AS, and QTL was designated QGw1.ccsu-1A. Received: 15 September 1999 / Accepted: 9 November 1999     

Single-trait,multi-locus and multi-trait GWAS using four different models for yield traits in bread wheat

Molecular Breeding - Rice (Oryza sativa L.) is a saline-alkali-sensitive crop. Saline-alkali environments can seriously affect the growth, development, and yield of rice. The mechanisms of salt...     

Amyloid Form of Ovalbumin Evokes Native Antigen-specific Immune Response in the Host: PROSPECTIVE IMMUNO-PROPHYLACTIC POTENTIAL

Amyloids are highly organized protein aggregates that arise from inappropriately folded versions of proteins or polypeptides under both physiological as well as simulated ambiences. Once thought to be irreversible assemblies, amyloids have begun to expose their more dynamic and reversible attributes depending upon the intrinsic properties of the precursor protein/peptide and experimental conditions such as temperature, pressure, structural modifications in proteins, or presence of chemicals in the reaction mixture. It has been repeatedly proposed that amyloids undergo transformation to the bioactive peptide/protein forms under specific conditions. In the present study, amyloids assembled from the model protein ovalbumin (OVA) were found to release the precursor protein in a slow and steady manner over an extended time period. Interestingly, the released OVA from amyloid depot was found to exhibit biophysical characteristics of native protein and reacted with native-OVA specific monoclonal as well as polyclonal antibodies. Moreover, antibodies generated upon immunization of OVA amyloidal aggregates or fibrils were found to recognize the native form of OVA. The study suggests that amyloids may act as depots for the native form of the protein and therefore can be exploited as vaccine candidates, where slow antigen release over extended time periods is a pre-requisite for the development of desired immune response.     

Use of methylation filtration and Cot fractionation for analysis of genome composition and comparative genomics in bread wheat

We investigated the compositional and structural differences in sequences derived from different fractions of wheat genomic DNA obtained using methylation filtration and Cot fractionation. Comparative analysis of these sequences revealed large compositional and structural variations in terms of GC content, different structural elements including repeat sequences (e.g., transposable elements and simple sequence repeats),protein coding genes, and non-coding RNA genes. A correlation between methylation status [determined on the basis of selective inclusion/exclusion in methylation-filtered (MF) library]of different repeat elements and expression level was observed. The expression levels were determined by comparing MF sequences with expressed sequence tags (ESTs) available in the public domain. Only a limited overlap among MF,high Cot (HC), and ESTs was observed, suggesting that these sequences may largely either represent the low-copy non-transcribed sequences or include genes with low expression levels. Thus, these results indicated a need to study MF and HC sequences along with ESTs to fully appreciate complexity of wheat gene space.     

Aufreinigung von Fumonisinen und ihren Hydrolysaten über ein neuartiges Festphasenextraktions-Material

Tests with various clean-up materials after optimisation of different parameters showed that the use of Oasis® material resulted in matrixless chromatograms in HPLC-FLD. The selectivity and detection limit of the method was improved by using LC-MS/MS as the detection system. Mean recovery was 100%, and no negative food matrix effects could be observed.