用免疫荧光单克隆抗体对脊髓灰质炎病毒抗原表位的分析

用间接免疫荧光与中和试验筛选出来的抗脊髓灰质炎2型与3型不同毒株的12个单克隆抗体,其中3个仅有免疫荧光活性,9个具有中和与免疫荧光活性。用免疫荧光活性的单克隆抗体进行试验,发现它们在识别特异性抗原表位方面与中和性单克隆抗体相似,显示出株特异的、几个毒株共同特异的或型特异的抗原表位。根据表位分布关系及特征,可以用来鉴别型内毒株的特征、毒株的抗原分析、抗原变异的研究以及疫苗相关病例的鉴别。所得结果与中和性单克抗隆抗体及T1-寡核苷酸指纹图谱分析一致。而免疫荧光单克隆抗体识别抗原表位的活性范围比中和性单克隆抗体更广。另外还发现某些兼有荧光与中和两活性的同一个单克隆抗体,用不同方法(IF与NT)进行试验时,与相同毒株出现不同表位反应,这点是值得引起注意需待进一步证实的重要问题。     

Isolation and Biochemical Characterization of a Neural Proteoglycan Expressing the L5 Carbohydrate Epitope

The monoclonal L5 antibody reacts with an N-glycosidically linked carbohydrate structure which is present on the neural cell adhesion molecule L1, neural chondroitin sulfate proteoglycans, and other not yet identified glycosylated proteins. Using this antibody, we isolated and characterized proteoglycans from adult mouse brain and cultured astrocytes biosynthetically labeled with Na2 35SO4 and a 3H-amino acid mixture. Our data suggest that the L5 proteoglycans of both sources are identical in their biochemical properties. The apparent molecular mass of the L5 proteoglycan is approximately 500 kDa. Digestion of the iodinated L5 proteoglycan from mouse brain and of the [35S]methionine-labeled L5 proteoglycan from cultured astrocytes with proteinase-free chondroitinases ABC and AC revealed three major core proteins with apparent molecular masses of approximately 380, 360, and 260 kDa. These represent molecularly distinct protein cores.     

Linkage mapping of genes for resistance to leaf,stem and stripe rusts and ω-secalins on the short arm of rye chromosome 1R

Summary The genes controlling resistance to three wheat rusts, viz., leaf rust (Lr26), stem rust (Sr31) and stripe or yellow rust (Yr9), and -secalins (Sec1), located on the short arm of rye chromosome 1R, were mapped with respect to each other and the centromere. Analysis of 214 seeds (or families derived from them) from testcrosses between a 1BL.1RS/1R heterozygote and Chinese Spring ditelocentric 1BL showed no recombination between the genes for resistance to the three rusts, suggesting very tight linkage or perhaps a single complex locus conferring resistance to the three rusts. The rust resistance genes were located 5.4 ± 1.7 cM from the Sec1 locus, which in turn was located 26.1 ± 4.3 cM from the centromere; the gene order being centromere — Sec1 — Lr26/Sr31/Yr9 — telomere. In a second test-cross, using a different 1BL.1RS translocation which had only stem rust resistance (SrR), the above gene order was confirmed despite a very large proportion of aneuploids (45.8%) among the progeny. Furthermore, a map distance of 16.0 ± 4.8 cM was estimated for SrR and the telomeric heterochromatin (C-band) on 1RS. These results suggest that a very small segment of 1RS chromatin is required to maintain resistance to all three wheat rusts. It should be possible but difficult to separate the rust resistance genes from the secalin gene(s), which are thought to contribute to dough stickiness of wheat-rye translocation lines carrying 1RS.     

Macrophytes and flood plain water dynamics in the River Danube ecotone research region (Austria)

In the ecotone research region of the Danube in Austria (Man-and-the-Biosphere (MaB)- project 5/21, Austrian Academy of Science) the macrophytes are one of the most important groups of organisms investigated. The species composition and the plant mass in hydrologically dynamic, and in predominantly stagnant sections of a system of lateral branches and sloughs were studied. This study showed that areas protected from frequent disturbance by floods had a greater number of species and higher biomass of aquatic macrophytes. Some species were shown to be evenly distributed throughout the branch system, whereas other species tended to form rare, but large singular patches. A set of new data elaboration techniques enabled us to describe the distribution pattern of the aquatic vegetation in this large branch system of the River Danube.     

Genetic analysis of two tomato mutants affected in the regulation of iron metabolism

Iron is one of the most important micronutrients for plants. Like other organisms, plants have developed active mechanisms for the acquisition of sufficient iron from the soil. Nevertheless, very little is known about the genetic mechanisms that control the active uptake. In tomato, two spontaneously derived mutants are available, which are defective in key steps that control this process. The recessive mutationchloronerva (chln) affects a gene which controls the synthesis of the non-protein amino acid nicotianamine (NA), a key component in the iron physiology of plants. The root system of the recessive mutantfer is unable to induce any of the characteristic responses to iron deficiency and iron uptake is thus completely blocked. We present a characterization of the double mutant, showing that thefer gene is epistatic over thechln gene and thus very likely to be one of the major genetic elements controlling iron physiology in tomato. In order to gain access to these two genes at the molecular level, both mutants were precisely mapped onto the high density RFLP map of tomato. Thechln gene is located on chromosome 1 and thefer gene is on chromosome 6 of tomato. Using this high-resolution map, a chromosome walk has been started to isolate thefer gene by map-based cloning. The isolation of thefer gene will provide new insights into the molecular mechanisms of iron uptake control in plants.     

A physical and genetic map of theCorynebacterium glutamicum ATCC 13032 chromosome

A combined physical and genetic map of theCorynebacterium glutamicum ATCC 13032 chromosome was constructed using pulsed-field gel electrophoresis (PFGE) and hybridizations with cloned gene probes. Total genomic DNA was digested with the meganucleasesSwaI (5-ATTTAAAT-3),PacI (5-TTAATTAA-3), andPmeI (5-GTTTAAAC-3) yielding 26, 27, and 23 fragments, respectively. The chromosomal restriction fragments were then separated by PFGE. By summing up the lengths of the fragments generated with each of the three enzymes, a genome size of 3082 +/- 20 kb was determined. To identify adjacentSwaI fragments, a genomic cosmid library ofC. glutamicum was screened for chromosomal inserts containingSwaI sites. Southern blots of the PFGE gels were hybridized with these linking clones to connect theSwaI fragments in their natural order. By this method, about 90% of the genome could be ordered into three contigs. Two of the remaining gaps were closed by cross-hybridization of blottedSwaI digests using as probesPacI andPmeI fragments isolated from PFGE gels. The last gap in the chromosomal map was closed by hybridization experiments using partialSwaI digestions, thereby proving the circularity of the chromosome. By hybridization of gene probes toSwaI fragments separated by PFGE about 30 genes, including rRNA operons, IS element and transposon insertions were localized on the physical map.     

Analysis of the Epitopes Recognized by Mouse Monoclonal Antibodies Directed to Yersinia enterocolitica Heat-Shock Protein 60

To determine amino acid sequences of the epitopes recognized by monoclonal antibodies (mAbs) 3C8 and 5C3 directed against Yersinia enterocolitica heat-shock protein (HSP60), a dot blot analysis was perfomed using synthesized peptides of Y. enterocolitica HSP60 such as peptides p316-342, p327-359, p340-366, p316-326, p316-321, p319-323, and p321-326 which represent positions of amino acids in Y. enterocolitica HSP60. The dot blot analysis revealed that 5C3 mAb reacted with p316-342, p316-326 and p321-326, and 3C8 mAb p316-342 and p316-326. These results indicate that the epitopes recognized by the mAbs were associated with eleven amino acids, Asp Leu Gly Gln Ala Lys Arg Val Val Ile Asn, of p316-326. The sequence homology between p316-326 of Y. enterocolitica HSP60 and the rest of the HSP60 family suggests that the five amino acids of Lys, Arg, Val, Ile and Asn, which are highly conserved in the HSP60 family, might be related with the epitope recognized by 3C8. In contrast, it was also demonstrated that three amino acids of Leu, Gly and Val, which are not well conserved in the HSP60 family, might be related to the epitope recognized by 5C3.     

Structures and contribution to the antigenicity of oligosaccharides of Japanese cedar (Cryptomeria japonica) pollen allergenCry j I: relationship between the structures and antigenic epitopes of plantN-linked complex-type glycans

The oligosaccharide structures ofCry j I, a major allergenic glycoprotein ofCryptomeria japonica (Japanese cedar, sugi), were analysed by 400 MHz¹H-NMR and two-dimensional sugar mapping analyses. The four major fractions comprised a series of biantennary complex type N-linked oligosaccharides that share a fucose/xylose-containing core and glucosamine branches including a novel structure with a nongalactosylated fucosylglucosamine branch.Rabbit polyclonal anti-Cry j I IgG antibodies cross-reacted with three different plant glycoproteins having the same or shorter N-linked oligosaccharides asCry j I. ELISA and ELISA inhibition studies with intact glycoproteins, glycopeptides and peptides indicated that both anti-Cry j I IgGs and anti-Sophora japonica bark lectin II (B-SJA-II) IgGs included oligosaccharide-specific antibodies with different specificities, and that the epitopic structures against anti-Cry j I IgGs include a branch containing 1–6 linked fucose and a core containing fucose/xylose, while those against anti-B-SJA-II IgGs include nonreducing terminal mannose residues. The cross-reactivities of human allergic sera to miraculin andClerodendron Trichotomum lectin (CTA) were low, and inhibition studies suggested that the oligosaccharides onCry j I contribute little or only conformationally to the reactivity of specific IgE antibodies.Abbreviations Cry j I a major allergenic glycoprotein ofCryptomeria japonica - B-SJA-II Sophora japonica bark lectin II - CTA Clerodendron trichotomum lectin - TFMS trifluoromethanesulfonic acid - HRP horseradish peroxidase