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PubMed1985

Development of IgE and IgG antibodies to food and inhalant allergens in children at risk of allergic disease.

In a prospective study of 92 children with at least one atopic parent, the development of the specific antibody responses to food and inhalant allergens during the first 5 years of life were assessed. By the radioallergosorbent test egg specific IgE antibody occurred in about 30% of the children with the mean peak concentration at 12 months. By the second year the prevalence of this antibody had increased whereas the mean concentration had decreased. Milk specific IgE antibody could not be shown in any subject, including four whose skin tests yielded positive results. Food specific IgG antibody was noted by antigen binding radioimmunoassays at 3 months in most children. These responses had peaked and began to fall by the fifth year. In contrast few children had detectable IgE or IgG antibody to inhalant allergens before the first 2 years of life. Both the concentration and prevalence of specific antibody, however, increased from the second to the fifth year and was greater in children whose skin tests yielded positive results. Breast feeding was associated with an increase in the prevalence of positive results from skin tests but was not associated with detectable IgE antibody to both food proteins, a lower concentration of IgG antibody to cows' milk, and was not associated with protection against the development of disease. A high level of exposure to dust mite was associated with an increased prevalence of positive results from skin tests to dust mite and appreciably higher antibody concentration. This study indicates differences in the humoral responses to food and inhalant allergens. Environmental factors appear to influence the development of these responses.

Aging↗
PubMed1974

The toxicity of certain cereal proteins in coeliac disease.

Gliadins from wheat, rye, and oats, and from wheat glutenin were digested with pepsin, trypsin, and pancreatin and the products (PTC digests) chromatographed on sulphopropyl (SP) Sephadex. Fractions eluted near neutral pH from wheat, rye, and oats gliadin digests all had very similar amino acid composition, although the oats fraction was higher in sulphur-containing amino acids. The major amino acids present in all were glutamine/glutamic acid and proline. The amount of fraction eluted near neutral pH from oats gliadin digest was about 13% of that eluted from digests of wheat and rye gliadins. Moreover, the yield of gliadin from oatmeal was only 0.5% compared with 2.4% and 2.8% from rye and wheat flours respectively. The amount of fraction eluted from wheat glutenin digest was about 70% of that obtained from wheat and rye gliadin digests. The fractions eluted near neutral pH from all protein digests were defectively digested by remission coeliac mucosa, and D-xylose excretion tests with the fraction from the wheat gliadin digest (fraction 9) indicated that it is harmful to subjects with coeliac disease, whereas the other fractions of this digest gave no such evidence. The results of the present work indicate that counterpart fractions to fraction 9 obtained from wheat glutenin and rye and oats gliadins may also be important in the aetiology of coeliac disease.

Adolescent↗
PubMed2002

Identification and chromosomal organization of two rye genome-specific RAPD products useful as introgression markers in wheat.

Two rye genome-specific random amplified polymorphic DNA (RAPD) markers were identified for detection of rye introgression in wheat. Both markers were amplified in all of the tested materials that contained rye chromatin such as rye, hexaploid triticale, wheat-rye addition lines, and wheat varieties with 1BL.1RS translocation. Two cloned markers, designated pSc10C and pSc20H, were 1012 bp and 1494 bp, respectively. Sequence analysis showed that both pSc10C and pSc20H fragments were related to retrotransposons, ubiquitously distributed in plant genomes. Using fluorescence in situ hybridization (FISH), probe pSc10C was shown to hybridize predominantly to the pericentromeric regions of all rye chromosomes, whereas probe pSc20H was dispersed throughout the rye genome except at telomeric regions and nucleolar organizing regions. The FISH patterns showed that the two markers should be useful to select or track all wheat-rye translocation lines derived from the whole arms of rye chromosomes, as well as to characterize the positions of the translocation breakpoints generated in the proximal and distal regions of rye arms.

Base Sequence↗
PubMed2003

The conversion of wheat RFLP probes into STS markers via the single-stranded conformation polymorphism technique.

We describe a flexible and general strategy for converting a wheat RFLP-based assay into a PCR-based sequence-tagged site (STS), and have applied it to derive markers for a powdery mildew resistance gene present in a wheat-rye translocation. The concept is based on deriving PCR primers that amplify all of the homoeoloci defined by a single-copy cDNA sequence, and separating the resulting mixture of homoeoamplicons via single-stranded conformation polymorphism (SSCP) gels, which are able to detect minor differences between related DNA sequences. After their separation, the individual homoeoamplicons were sequenced and these were used to define nucleotide polymorphisms that could be exploited to design locus-specific PCR primers. In one case, we were able to demonstrate that the assay was allele specific.

Base Sequence↗
PubMed2004

Genetic and physical mapping of homoeologous recombination points involving wheat chromosome 2B and rye chromosome 2R.

Wide hybrids have been used in generating genetic maps of many plant species. In this study, genetic and physical mapping was performed on ph1b-induced recombinants of rye chromosome 2R in wheat (Triticum aestivum L.). All recombinants were single breakpoint translocations. Recombination 2RS-2BS was absent from the terminal and the pericentric regions and was distributed randomly along an intercalary segment covering approximately 65% of the arm's length. Such a distribution probably resulted from structural differences at the telomeres of 2RS and wheat 2BS arm that disrupted telomeric initiation of pairing. Recombination 2RL-2BL was confined to the terminal 25% of the arm's length. A genetic map of homoeologous recombination 2R-2B was generated using relative recombination frequencies and aligned with maps of chromosomes 2B and 2R based on homologous recombination. The alignment of the short arms showed a shift of homoeologous recombination toward the centromere. On the long arms, the distribution of homoeologous recombination was the same as that of homologous recombination in the distal halves of the maps, but the absence of multiple crossovers in homoeologous recombination eliminated the proximal half of the map. The results confirm that homoeologous recombination in wheat is based on single exchanges per arm, indicate that the distribution of these single homoeologous exchanges is similar to the distribution of the first (distal) crossovers in homologues, and suggest that successive crossovers in an arm generate specific portions of genetic maps. A difference in the distribution of recombination between the short and long arms indicates that the distal crossover localization in wheat is not dictated by a restricted distribution of DNA sequences capable of recombination but by the pattern of pairing initiation, and that can be affected by structural differences. Restriction of homoeologous recombination to single crossovers in the distal part of the genetic map complicates chromosome engineering efforts targeting genes in the proximal map regions.

Chromosome Mapping↗
PubMed2003

DNA, chromosomes, and in situ hybridization.

In situ hybridization is a powerful and unique technique that correlates molecular information of a DNA sequence with its physical location along chromosomes and genomes. It thus provides valuable information about physical map position of sequences and often is the only means to determine abundance and distribution of repetitive sequences making up the majority of most genomes. Repeated DNA sequences, composed of units of a few to a thousand base pairs in size, occur in blocks (tandem or satellite repeats) or are dispersed (including transposable elements) throughout the genome. They are often the most variable components of a genome, often being species and, occasionally, chromosome specific. Their variability arises through amplification, diversification and dispersion, as well as homogenization and loss; there is a remarkable correlation of molecular sequence features with chromosomal organization including the length of repeat units, their higher order structures, chromosomal locations, and dispersion mechanisms. Our understanding of the structure, function, organization, and evolution of genomes and their evolving repetitive components enabled many new cytogenetic applications to both medicine and agriculture, particularly in diagnosis and plant breeding.

Animals↗
PubMed2005

Molecular and cytological characterization of genomic variability in hexaploid wheat 'Lindström'.

'Lindström' wheat (AABBDD+rye B chromosomes) was used to study the effects of alien chromatin introgressed into a wheat genetic background, subjecting the wheat genome to a new and transient allopolyploidisation episode. Using this experimental material, we have previously demonstrated that no large-scale chromosomal translocations occurred as a result of the genomic constitution of the addition line. However, we have shown that the presence of a number of rye B chromosomes is associated with changes in the interphase organization and expression patterns of wheat rDNA loci. We have now extended our studies to focus on a further characterization of 'Lindström' 5S rDNA loci and also on high molecular weight glutenin subunit (HMW-GS) patterns. In the process, we have uncovered an unusually large variant of the 5S rDNA locus on wheat chromosome 1B (not to be confused with rye B chromosomes) and 2 novel HMW glutenin y-type alleles. These changes are not directly related to variation in rye B chromosome number in the present material, but the fact that a new, and still segregating, 1Dy HMW-GS gene was identified indicates a recent timescale for its origin. Strikingly, the 'Lindström' 5S rDNA 1B locus integrates a unit sharing 94% homology with a rye 5S rDNA sequence, suggesting the possibility that the wheat locus was colonized by highly homologous rye sequences during the breeding of 'Lindström', when the rye and wheat genomes were together, albeit briefly, in the same nucleus.

Base Sequence↗
PubMed2005

Mapping of QTLs for androgenetic response based on a molecular genetic map of x Triticosecale Wittmack.

Quantitative trait loci (QTLs) for androgenetic response were mapped in a doubled haploid (DH) population derived from the F1 hybrid of 2 unrelated varieties of triticale, 'Torote' and 'Presto'. A molecular marker linkage map of this cross was previously constructed using 73 DH lines. This map contains 356 markers (18 random amplified 5 polymorphic DNA, 40 random amplified microsatellite polymorphics, 276 amplified fragment length polymorphisms, and 22 simple sequence repeats) and was used for QTL analysis. The genome was well covered, and of the markers analysed, 336 were located in 21 linkage groups (81.9%) identified using SSR markers. The map covered a total length of 2465.4 cM with an average of 1 marker for each 6.9 cM. The distribution of the markers was not homogeneous across the 3 genomes, with 50.7% detected in the R genome. Several QTLs were found for the following variables related to the androgenetic response: number of embryos/100 anthers; plants regenerated from 100 embryos; number of green plants/total number of plants; and number of green plants/1000 anthers. Two were detected on chromosome 6B and 4R, which together had a 30% total influence on the induction of embryos. Another was found on 6B and on the unidentified LG1; these influenced the production of total plants from haploid embryo cultures. One QTL on chromosome 3R determined the photosynthetic viability of the haploid plantlets regenerated from microspores. Other QTLs were found on chromosomes 1B, 1R, 4R, and 7R, which helped the control of the final androgenetic response (the number of plantlets obtained for every 1000 anthers cultured).

Chromosome Mapping↗
PubMed1993

An optimized fluorescence in situ hybridization procedure for detecting rye chromosomes in wheat.

In situ hybridization with an interspersed repeat clone from rye, pSc119, was shown to be useful for detecting rye chromosomes introduced into wheat. However, since pSc119 also shows strong hybridization to a few sites in certain wheat chromosomes, small rye chromosome segments added to wheat may be difficult to detect. In this study, detection of rye chromosomes present in triticale and triticale X wheat hybrids was accomplished with the use of a subfragment from pSc119 (pSc119.1) whose sequence is dispersed throughout the rye chromosomes and only weakly cross-hybridizes to a few telomeric and centromeric regions of wheat. The in situ hybridization conditions were optimized to readily distinguish rye chromosomes from wheat chromosomes without the need for intensive analysis of hybridization patterns. Rye chromosomes were readily detected using fluorescence in situ hybridization. Fluorescence detection provided increased sensitivity over enzymatic detection and allowed signals to be amplified with repeated use of biotinylated anti-avidin antibody and avidin-FITC. Detection of rye chromatin was further optimized by doubling the probe concentration. Finally, double exposure photography of the same cell with two different filters provided another means to further increase the contrast between rye and wheat chromosomes.

Base Sequence↗
PubMed1994

Detection of rye chromosome 2R using the polymerase chain reaction and sequence-specific DNA primers.

Sequences derived from a rye gamma secalin gene were used as primers in polymerase chain reactions using DNA obtained from a series of wheat and triticale genetic stocks. A 473-bp fragment, the predicted size based on the distance between the selected primers, was found only in rye, triticales, and wheat lines carrying rye chromosome 2RS. Use of triticale lines with various wheat chromosome substitutions confirmed the chromosomal origin of the rye-specific marker. The presence of the 473-bp PCR product was always associated with the production of 75K secalins in grain samples. Thus, the primer sequences, and the clone of origin (pSC503), were both derived from the SEC-2 locus of rye chromosome 2RS.

Base Sequence↗
PubMed1994

Amplification of DNA sequences in wheat and its relatives: the Dgas44 and R350 families of repetitive sequences.

The sequence of a Triticum tauschii genomic clone representing a family of D-genome amplified DNA sequences, designated Dgas44, is reported. The Dgas44 sequence occurs on all chromosomes of the D genome of wheat, Triticum aestivum, and in situ hybridization revealed it to be evenly dispersed on all seven chromosome pairs. An internal HindIII fragment of Dgas44, designated Dgas44-3, defines the highly amplified region that is specific to the D genome. The polymerase chain reaction was used to amplify a 236-bp fragment within Dgas44-3 from chromosomes 1D, 2D, 3D, 4D, 5D, and 7D, and identical copies of this region of the Dgas44-3 sequence were found among the isolates from each of the chromosomes. The Dgas44-3 sequence population from specific chromosomes differed on average by 0.22% from the original Dgas44 sequence. The Dgas44 sequence was found to differentiate between the D genome present in T. aestivum, T. tauschii, hexaploid T. crassum, T. cylindricum, T. ventricosum, in which the sequence was present in a highly amplified form and T. juvenale, T. syriacum, and tetraploid T. crassum where the sequence family was difficult to detect. Another class of amplified sequences previously considered to be rye "specific." R350, was isolated from tetraploid wheat and its dispersed distribution on chromosomes was similar to the Dgas44 family in T. tauschii. In contrast with the Dgas44 sequence family, genome specificity for the remnant R350 sequence family was not evident since it was present on all wheat chromosomes.

Base Sequence↗
PubMed1996

A PCR assay for detection of a 2RL.2BS wheat-rye chromosome translocation.

A 2RL.2BS wheat-rye translocation, present in the wheat germplasm line Hamlet, carries a gene for resistance to Hessian fly biotype L, one of the most virulent biotypes presently encountered in wheat production environments. Unlike several other wheat-rye chromosome translocations common in wheat breeding programs, 2RL lacks genes encoding storage proteins or other easily selected markers. Oligonucleotide primers synthesized from published sequences derived from the R173 family of moderately repetitive rye DNA were used in the DNA polymerase chain reaction (PCR) to identify specific markers for 2RL. The same primers, when used with DNA extracted from additional wheat-rye translocation lines of importance to the wheat breeding community, gave distinctive PCR products for each genotype. The single primer pair, PAWS5 and PAWS6, may, therefore, have wide applicability for the identification of wheat-rye chromosomal translocations presently encountered in wheat breeding populations.

Base Sequence↗
PubMed1996

Chromosomal location of three wheat sequences with homology to pollen allergen encoding, DNA replication regulating, and DNA (cytosine-5)-methyltransferase genes in wheat and rye.

Three wheat sequences, shown to be homologous to pollen allergen encoding, DNA replication regulating, and DNA (cytosine-5)-methyltransferase genes were localized on chromosomes using nullisomic-tetrasomic wheat ('Chinese Spring') and wheat-rye ('Chinese Spring'/'Imperial') addition lines. Whereas the loci for the pollen allergen encoding sequence (Tri a III) were shown to be located on homoeologous group 4, the DNA replication regulating (Rep) and DNA (cytosine-5)-methyltransferase (Mtase) genes were located to homoeologous groups 1 and 7, respectively, of Triticeae. Chromosomal rearrangements in wheat and rye relative to each other are discussed.

Allergens↗
PubMed1979

Effect of irradiance upon the population of Pseudomonas coronafaciens in leaves and symptom expression of halo blight of rye.

The toxin-induced chlorosis caused by Pseudomonas coronafaciens is influenced by irradiance. Three levels of irradiance caused differences in symptom expression but did not affect the rate of increase or final population of viable cells of P. coronafaciens in rye leaves. Distinct and faint halo blight symptoms appeared in 3--4 days in full light (1425 microW cm-2), and 58% shade (598 microW cm-2) respectively. No symptoms or only faint symptoms appeared after 7 days at 86% shade (202 microW cm-2). When plants kept in 58 and 86% shade were moved to full light 5 days after inoculation, lesion size and chlorosis increased rapidly during the next 2 days. On the 7th day after inoculation, the size of lesions from the 58 and 86% shade treatments exceeded those in full light by 2.5 and 5 times, respectively. A chlorosis index based on lesion size and chlorophyll loss within lesions also reflected this trend although chlorophyll loss was greater in lesions in full light for 7 days. Conditions of low irradiance such as that caused by overcast weather and (or) a dense leaf canopy followed by bright sunshine can cause greater losses from halo blight than a continuous period of high irradiance. Sympton expression may be masked by low irradiance but increase of inoculum is not impaired. Although increased light enhances chlorosis, toxin diffusion or production may be reduced.

Chlorophyll↗
PubMed1976

The selective extraction of histones from rye chromatin.

The selective extraction of histones from rye chromatin was studied using three different methods. Extractions with NaCl-phosphate buffers at pH5.5 gave results similar to those already obtained with other types of chromatin. Histone H1 was selectively extracted with 0.6 M NaCl-0.001 M PO4, while the selectivity of dissociation of the other fractions was reduced at higher NaCl concentrations. The use of phosphate-urea buffers at pH5.5 also revealed that the histones were dissociated at the same concentrations as were calf thymus histones. Histone H1 was extracted with 0.5 M PO4-1 M urea; H1, H2A, and H2B were extracted with 0.8 M PO4-2 M urea; and all histones were removed with 0.8 M PO4-5.3 M urea. It was, however, observed that the dissociated rye histones H2A and H2B were unstable in these buffers. This instability was maximum in the presence of 3 M urea, where both histones were absent from the extracted proteins and the residual nucleoproteins. Finally, a solution of 30% ethanol-0.35 M NaCl-6 M urea produced a rye nucleoprotein fraction containing only histone H1.

Animals↗
PubMed1976

Chloroplastic proteins of wheat and rye grown at warm and cold-hardening temperatures.

Soluble proteins and membrane polypeptides were separated from chloroplasts isolated intact from a cultivar each of spring wheat, winter wheat, and more freeze-resistant rye, and changes in them associated with cold hardening were detected by means of polyacrylamide gel electrophoresis. No drastic changes in chloroplast membrane polypeptides occurred during growth at low temperatures in the three cultivars. However, subtle changes were evident in the soluble chloroplast protein fraction. In this fraction at least one varietal difference was discernable, yet all cultivars produced a new protein band of lowest mobility during growth at low temperatures. After the preparations were fractionated by Sephadex G-50 all unhardened plant material displayed two peaks in the region of the fraction I protein band, whereas all cold-hardening material displayed one peak. A different band of soluble protein was present only after cold hardening in Kharkov whear and Puma rye, and was not present in extracts from the cold-grown spring wheat.

Chloroplasts↗
PubMed1978

Evidence for an in vivo conformational change in ribulose bisphosphate carboxylase-oxygenase from Puma rye during cold adaptation.

Ribulose bisphosphate carboxylase-oxygenase (RUBPCase) from leaves of cold-hardened and unhardened Puma rye was purified by gel filtration and ion exchange chromatography. Chemical properties that might be associated with a previously demonstrated difference in molecular charge of purified RUBPCase from cold-hardened and unhardened Puma rye were investigated. Amino acid analyses indicated no significant differences in amino acid composition or average hydrophobicity per residue. Enzymes from hardened and unhardened rye were reversibly cold inactivated at 0 degrees C. However, the former was more stable at this temperature than the latter. Titration with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) indicated the presence of fast and slow titrating sulfhydryl groups in both enzyme preparations but there were 50% fewer SH groups titrated in the enzyme from hardened rye in 30 min that in the enzyme from unhardened rye. Activation of both enzymes by HCO-3 enhanced the reactivity of sulfhydryl groups to titration with DTNB. According to the kinetics of the slow titrating SH groups, the enzyme from hardened rye was less susceptible to denaturation by sodium dodecyl sulfate than was the same enzyme from unhardened rye. It is concluded that the tertiary structure of RUBPCase from Puma rye is affected during low-temperature adaptation.

Adaptation, Physiological↗