Diagnosis of alpha 1-antitrypsin deficiency by enzymatic amplification of human genomic DNA and direct sequencing of polymerase chain reaction products.

We have compared sequencing of cloned "polymerase chain reaction" (PCR) products and the direct sequencing of PCR products in the examination of individuals from six families affected with alpha 1-antitrypsin (AAT) deficiency. In families where paternity was in question we confirmed consanguinity by DNA fingerprinting using a panel of locus-specific minisatellite probes. We demonstrate that direct sequencing of PCR amplification products is the method of choice for the absolutely specific diagnosis of AAT deficiency and can distinguish normals, heterozygotes and homozygotes in a single, rapid and facile assay. Furthermore, we demonstrate the reproducibility of the PCR and a rapid DNA isolation procedure. We have also shown that two loci can be simultaneously amplified and that the PCR product from each locus can be independently examined by direct DNA sequencing.     

The structure and phenology of a moist deciduous forest in the Central Indian Highlands

The vegetation structure and phenology of a 74.5 ha block of moist deeiduous forest in Kanha Tiger Reserve, Madhya Pradesh, India were investigated during primatological fieldwork. Within a grid of 0.25 ha quadrats all 9935 trees (woody plants > 2 m tall), of 63 species, were enumerated and their girth measured. The forest was dominated by the dipterocarp sal (Shorea robusta). Most species were rare and showed clumped distributions within the sea of sal whilst the dispersion of five large canopy trees could not be distinguished from random.The phenology of 215 trees, of 61 species, was monitored over 14 months using an index of phytophase abundance. Leaf renewal was highly synchronous between and within species, mostly occurring between February and June. One species was evergreen, 5 were semi-evergreen and 55 were deciduous. Flowering generally occurred between leaf fall and flush whilst fruiting peaked late in the hot weather and early monsoon.     

Enzyme-linked immunosorbent assays for the measurement of blood group A and B glycosyltransferase activities

ELISA assays have been developed for (1–3)N-acetylgalactosaminyltransferase (blood group A transferase) and (1–3)galactosyltransferase (blood group B transferase) activities. In these assays, microtitre plates coated with the bovine serum albumin conjugate of a synthetic Fuc1–2Gal-R acceptor substrate are incubated with the appropriate nucleotide donor (UDP-GalNAc or UDP-Gal) and human serum as the enzyme source. The resulting trisaccharide products Fuc1–2(GalNAc1–3)Gal-R-BSA or Fuc1–2(Gal1–3)Gal-R-BSA are detected and quantified with monoclonal antibodies selected not to cross-react with the substrate structure. With less than a microliter of human serum, product formation is proportional to enzyme concentration and to time of incubation of up to 90 min.     

A minority of 46,XX true hermaphrodites are positive for the Y-DNA sequence including SRY

Summary A total of 30 cases of 46,XX true hermaphroditism was analysed for Y-DNA sequences including the recently cloned gene for male testis-determination SRY. In 3 cases, a portion of the Y chromosome including SRY was present and, in 2 cases, was localised, to Xp22 by in situ hybridisation. Since previous studies have shown that the majority of XX males are generated by an X-Y chromosomal interchange, the Xp22 position of the Yp material suggests that certain cases of hermaphroditism can arise by the same meiotic event. The phenotype in the 3 SRY-positive cases may be caused by X-inactivation resulting in somatic mosaicism of testis-determining factor expression giving rise to both testicular and ovarian tissues. Autosomal or X-linked mutation(s) elsewhere in the sex-determining pathway may explain the phenotype observed in the remaining 27 SRY-negative cases.     

A 2.6 kb intron separates the signal peptide coding sequence of an anther-specific protein from the rest of the gene in sunflower

Summary Metabolism of sulfonylurea herbicides by Streptomyces griseolus ATCC 11796 is carried out via two cytochromes P-450, P-450_SU1 and P-450_SU2. Mutants of S. griseolus, selected by their reduced ability to metabolize a fluorescent sulfonylurea, do not synthesize cytochrome P-450_SU1 when grown in the presence of sulfonylureas. Genetic evidence indicated that this phenotype was the result of a deletion of > 15 kb of DNA, including the structural genes for cytochrome P-450_SU1 and an associated ferredoxin Fd-1 (suaC and suaB, respectively). In the absence of this monooxygenase system, the mutants described here respond to the presence of sulfonylureas or phenobarbital in the growth medium with the expression of only the suhC,B gene products (cytochrome P-450_SU2 and Fd-2), previously observed only as minor components in wild-type cells treated with sulfonylurea. These strains have enabled an analysis of sulfonylurea metabolism mediated by cytochrome P-450_SU2 in the absence of P-450_SU1, yielding an in vivo delineation of the roles of the two different cytochrome P-450 systems in herbicide metabolism by S. griseolus.     

Gene expression in nematode-infected plant roots

Summary A major pathogen of potato plants (Solanum tuberosum) is the potato cyst nematode (Globodera spp.), which induces localized redifferentiation of a limited number of host cells to form a specialized feeding-site termed the syncytium. A novel strategy utilizing the polymerase chain reaction (PCR) was employed to construct a cDNA library from dissected potato roots highly enriched in syncytial material. The library was differentially screened with cDNA probes derived from the infected root tissue from a compatible interaction and from healthy root tissue. Characterization of one gene identified by the library screen indicated an expression pattern that correlated with events in the immediate vicinity of the pathogen after syncytial establishment. The strategy for library construction and screening could be applicable to the study of gene expression in any plant-pathogen interaction in which the limited supply of cells at the interface of the two organisms precludes a more traditional approach.     

Chloroplast Structure and Function Is Altered in the NCS2 Maize Mitochondrial Mutant

The nonchromosomal stripe 2 (NCS2) mutant of maize (Zea mays L.) has a DNA rearrangement in the mitochondrial genome that segregates with the abnormal growth phenotype. Yet, the NCS2 characteristic phenotype includes striped sectors of pale-green tissue on the leaves. This suggests a chloroplast abnormality. To characterize the chloroplasts present in the mutant sectors, we examined the chloroplast structure by electron microscopy, chloroplast function by radiolabeled carbon dioxide fixation and fluorescence induction kinetics, and thylakoid protein composition by polyacrylamide gel electrophoresis. The data from these analyses suggest abnormal or prematurely arrested chloroplast development. Deleterious effects of the NCS2 mutant mitochondria upon the cells of the leaf include structural and functional alterations in the both the bundle sheath and mesophyll chloroplasts.     

Idioms of distress: Somatic responses to distress in everyday life

In-depth interviews were conducted among 50 subjects residing in the industrial town of Newcastle, Australia. Half of these subjects were from the general population and half were currently seeking counselling for personal/family problems. None of the subjects were receiving any medical care at the time of interview, though seven had done so during the episode of distress they were discussing. The study shows that while the subjects psychologized their problems, members of both groups tended to somatize at a rate proportional to the level of distress. Subjects were unaware of any relationship between the distress they were experiencing and their physical complaints. The results of this study support previous research which argues that those experiencing distress and those who tend to introspect are also those who are likely to amplify somatic symptoms. At the same time these results depart from findings in the United States which suggest that in the West, people learn to express social and personal distress in psychological terms,, thereby reducing the level of somatization. Though not representative of the population as a whole, the findings raised questions warranting further study.     

Lymphoma-vesicle interactions: vesicle adsorption, membrane fragmentation, and intermembrane protein transfer

Sonicated dimyristoylphosphatidylcholine vesicles interact with cultured murine lymphoma (BL/VL3) to generate complexes of vesicle and cell membrane components. Cell-free supernatants harvested after cell-vesicle incubations contain three distinct lipid species that can be separated by density gradient centrifugation. Analysis of protein and lipid composition and assays for cell and vesicle lumen contents reveal that the densest of the three lipid species comprises sealed plasma membrane fragments complexed with vesicles, while the least dense species is indistinguishable from pure phospholipid vesicles. The third, intermediate density species consists of topologically intact vesicles with associated plasma membrane proteins but without detectable cell lipids or cytoplasmic components. The membrane fragmentation and cell-to-vesicle protein transfer observed during lymphoma-vesicle incubations are examined as functions of cell and vesicle concentrations and incubation time.