Cytological and molecular characterization of a chromosome interchange and addition lines in Cadet involving chromosome 5B of wheat and 6Ag of Lophopyrum ponticum

Efforts to transfer wheat curl mite (Eriophyes tulipae Keifer) resistance from Lophopyrum ponticum 10X (Podb.) Love to bread wheat (Triticum aestivum L.) have resulted in the production of a number of cytogenetic stocks, including an addition line of 6Ag, a ditelo addition line, and a wheat-Lophopyrum translocation line. Characterization of these lines with C-banding, in situ hybridization with a Lophopyrum species-specific repetitive DNA probe (pLeUCD2), and Southern blotting with pLeUCD2 and a 5S ribosomal DNA probe (pScT7) confirmed that the distal portion of the short arm of 6Ag was translocated onto the distal portion of 5BS (5BL. 5BS-6AgS). It was also determined that the ditelo addition was an acrocentric chromosome of 6AgS.     

Isolation of carp cDNA clones,representing developmentally-regulated genes,using a subtractive-hybridization strategy

A subtractive-hybridization technique, combined with differential screenings and subsequent whole mount in situ hybridization (ISH) reactions, was used to isolate novel cDNA clones representing developmentally-regulated genes of carp. Small-scale differential screenings of an oocyte and a segmentation-stage cDNA library using oocyte-specific and segmentation stage-specific enriched probes, yielded 75 positive clones. ISH screening showed that 65% (15) of the oocyte-stage clones and 50% (26) of the segmentation-stage clones were indeed stage-specific. Partial sequence analysis suggests that approximately 65% of the 41 stage-specific clones represent novel genes. In addition, an Otxl clone was isolated. Two novel clones and the Otxl clone are of special interest for developmental studies. The clones represent genes that are locally expressed during embryonic development. The expression patterns of Otxl and one of the novel clones suggest functions in specification of the anterior-posterior axis. The three clones provide molecular markers for the study of gastrulation and the patterning of the a-p axis in teleosts.     

有机盐对基因启动子的σ因子选择性的影响

用σ￣（７０）或σ￣（３８）和核心酶（Ｅ）组成的大肠杆菌ＲＮＡ聚合酶全酶（Ｅσ）对四种启动子进行了体外转录。结果表明：ｌａｃＵＶ５,ｒｐｌＪ主要被Ｅσ￣（７０）识别,ｋａｔＥ主要被Ｅσ￣（３８）识别,ｆｉｃ在低盐浓度时被Ｅσ￣（７０）识别,在高浓度盐时被Ｅσ￣（３８）识别;Ｅσ￣（３８）转录特异性启动子所需酶量大于Ｅσ￣（７０）转录特异性启动子的酶量;在含有分别对σ￣（７０）或σ￣（３８）亲和性大的混和启动子的体外转录中,启动子之间不存在干扰和竞争,转录水平与启动子浓度成正相关。     

Isolation,characterization and application of a species-specific repeated sequence from Haynaldia villosa

A species-specific repeated sequence, pHvNAU62, was cloned from Haynaldia villosa, a wheat relative of great importance. It strongly hybridized to H. villosa, but not to wheat. In situ hybridization localized this sequence to six of seven H. villosa chromosome pairs in telomeric or sub-telomeric regions. Southern hybridization to whea-H. villosa addition lines showed that chromosomes 1V through 6V gave strong signals in ladders while chromosome 7V escaped detection. In addition to H. villosa, several Triticeae species were identified for a high abundance of the pHvNAU62 repeated sequence, among which Thinopyrum bassarabicum and Leymus racemosus produced the strongest signals. Sequence analysis indicated that the cloned fragment was 292 bp long, being AT rich (61%), and showed 67% homology of pSc7235, a rye repeated sequence. Isochizomer analysis suggested that the present repeated sequence was heavily methylated at the cytosine of the CpG dimer in the genome of H. villosa.It was also demonstrated that pHvNAU62 is useful in tagging the introduced 6VS chromosome arm, which confers a resistance gene to wheat powdery mildew, in the segregating generations.     

The UDP-Galactose:Ceramide Galactosyltransferase: Expression Pattern in Oligodendrocytes and Schwann Cells During Myelination and Substrate Preference for Hydroxyceramide

Abstract: Galactosylceramide ("galactocerebroside"; GalC) is a major glycolipid in the myelin sheath of the CNS and the PNS. The enzyme UDP-galactose:ceramide galactosyltransferase (CGalT) catalyzes the final step of the synthesis of GalC: the transfer of galactose to ceramide. By a differential screening approach, we have isolated a cDNA, the sequence of which is identical to the recently isolated cDNA clones for CGalT. By northern analysis and in situ hybridization we demonstrated that CGalT mRNA is expressed at birth in oligodendrocytes and Schwann cells, an expression pattern corresponding to the onset of myelination. In addition to the high expression levels of CGalT in oligodendrocytes and Schwann cells, in situ hybridization also showed expression in subtypes of neurons in spinal cord, cerebellum, and brainstem in the adult CNS, but at a much lower level than in oligodendrocytes. Expression of CGalT in COS cells demonstrated that CGalT has a preference for hydroxyceramide as a substrate. CGalT-expressing COS cells synthesize and transport GalC to their cell surface as shown by immunofluorescence and by lipid analysis of living cells. Our results suggested that the CGalT specifically uses hydroxyceramide for the synthesis of GalC and that separate (co)enzymes are not needed.     

Cell migration within the embryonic limb primordium of Drosophila as revealed by a novel fluorescence method to visualize mRNA and protein

 We report a new technique using fluorescent probes to detect a mRNA and a protein simultaneously in the Drosophila embryo. For in situ hybridization, 3-hydroxy-N-2′-biphenyl-2-naphthalenecarboxamide phosphate ester (HNPP)/Fast Red TR was used as a fluorescent substrate for alkaline phosphatase. It was possible to compare protein and mRNA expression on a cell by cell basis with a laser scanning confocal microscope. We applied this technique to analyse the dynamics of Distal-less (Dll) enhancer activity in the thoracic limb primordium in the early Drosophila embryo. We stained embryos bearing the Dll early enhancer (Dll-304) fused to the Escherichia coli lacZ gene. LacZ mRNA was detectable in the ventral region of the limb primordium, and β-galactosidase protein in the dorsal region. In the middle, both mRNA and protein were detectable. These results suggest that the Dll enhancer is activated in the ventral region of the limb primordium and that Dll-positive cells migrate from a ventral position to a dorsal one within a single limb primordium. Received: 7 April 1997 / Accepted: 15 May 1997     

Influence of muscle activity on the forces in the femur: An in vivo study

Experiments were performed on two patients with custom-made instrumented massive proximal femoral prostheses implanted after tumour resection. In vivo axial forces transmitted along the prostheses were telemetered during level walking, single- and double-leg stance, and isometric exercises of the hip muscles. These activities varied the lever arms available to the external loads: minimum for double-leg stance and maximum for hip isometric exercises. Kinematic, force plate, EMG and telemetered force data were recorded simultaneously. The force magnification ration (FMR; the ratio of the telemetered axial force to the external force) was calculated. The FMRs ranged from 1.3 (during double-leg stance) to 29.8 (during abductors test), indicating that a major part of the axial force in the long bones is a response to muscle activity, the strength of which depends on the lever arms available to the external loads. From these results, it was shown that the bulk of the bending moment along limbs is transmitted by a combination of tensile forces in muscles and compressive forces in bones, so moments transmitted by the bones are smaller than the limb moments. It was concluded that appropriate simulation of muscle forces is important in experimental or theoretical studies of load transmission along bones.     

Inositol 1,4,5-Trisphosphate 3-Kinase mRNA: High Levels in the Rat Hippocampal CA1 Pyramidal and Dentate Gyrus Granule Cells and in Cerebellar Purkinje Cells

The distribution of inositol 1,4,5-trisphosphate (InsP3) 3-kinase mRNA in the rat brain is reported using oligonucleotides based on a cDNA clone sequence that encodes rat brain InsP3 3-kinase and the in situ hybridization technique. Moderate levels were found in CA2-4 pyramidal neurons, in the cortex, and in the striatum. The cerebellar granule cells, thalamus, hypothalamus, brainstem, spinal cord, and white matter tracts were almost negative. The levels of InsP3 3-kinase mRNA were highest in the hippocampal CA1 pyramidal neurons, granule cells of the dentate gyrus, and cerebellar Purkinje cells. These results contrast with the lower concentration of the InsP3 receptor already reported in the hippocampus versus the Purkinje cells and suggest a special role for inositol 1,3,4,5-tetrakisphosphate in Ammon's horn.     

Uptake, Metabolism, and Release of N-[3H]-Acetylaspartylglutamate by the Avian Retina

N-Acetylaspartylglutamate (NAAG) is a nervous system-specific dipeptide that is released from retinal neurons on depolarization. In the present study, extracellular metabolism, uptake, and release of [3H]NAAG were examined in the chick retina. After in vitro incubation with NAAG radiolabeled in the glutamate moiety, [3H]glutamate and [3H]NAAG increased in retinal cells through time- and temperature-dependent processes, which were reduced in the absence of extracellular sodium. Coincubation of cells with [3H]NAAG and aspartylglutamate or phosphate resulted in the decreased extracellular appearance of [3H]glutamate, produced by hydrolysis of radiolabeled NAAG, and a consequent increased availability of [3H]NAAG for transport into the retinal cells. When this tissue was incubated with radiolabeled NAAG, glutamate, glutamine, or aspartate under similar conditions, only [3H]NAAG served as a significant source for the appearance of intracellular [3H]NAAG. These data support the conclusion that [3H]NAAG can be transported into retinal cells, whereas [3H]glutamate transport is the predominant process after release of this amino acid from NAAG by extracellular peptidase activities. After uptake, [3H]NAAG entered a cellular pool, from which the peptide was secreted under depolarizing conditions and in a calcium-dependent manner.