Rapid acquisition of three-dimensional triple-resonance experiments using pulsed field gradient techniques

Summary A rapid method for recording three-dimensional triple-resonance experiments utilising pulsed field gradient techniques is proposed, and applied to the HNCO experiment. In order to optimise the sensitivity of the method, a short phase cycle is used in conjunction with the pulsed field gradients to select the desired coherence transfer pathway. The method is demonstrated for the HU protein.     

Identification and sequence of a Na+-linked gene from the marine bacterium Alteromonas haloplanktis which functionally complements the dagA gene of Escherichia coli

A 4.0 kb fragment from a plasmid genomic DNA library of the marine bacterium Alteromonas haloplanktis ATCC 19855 was found in the presence of Na+ to complement the dagA gene of Escherichia coli. We have completely sequenced this fragment and the position of the Na(+)-linked D-alanine glycine permease gene (dagA) on the fragment has been determined by complementation. The predicted carrier protein consists of 542 amino acid residues (M(r) 58,955). Its hydropathy profile suggests it is composed of eight transmembrane segments with a long hydrophilic region between segments six and seven. Significant similarity has been found between this Na(+)-linked permease and the Na+/proline permeases of E. coli and Salmonella typhimurium and the human and rabbit intestinal Na+/glucose cotransporters.     

Oestrogen receptor gene structure and function in breast cancer

The mechanisms underlying loss of oestrogen responsiveness in breast cancer are not well-defined. Potential mechanisms include loss of receptor expression, alterations in the oestrogen receptor (ER) gene producing proteins with abnormal function, or changes to receptor-dependent or -independent pathways controlling cell proliferation. Examination by Southern analysis of the ER gene in a series of ER-negative and -positive breast tumour biopsies failed to provide evidence of gross rearrangements and in only only one of thirty seven tumour DNA samples was significant gene amplification observed. No restriction fragment length polymorphisms were detected for the restriction enzymes EcoRI, Pst I or Hind III. Methylation of the ER gene as assessed by Hpa II and Msp I restriction enzyme digests varied between tumours but the degree of methylation was not correlated with levels of expression of the receptor protein. Similar findings applied in a series of ER-negative and -positive breast cancer cell lines and clonal lines of MCF-7 cells, which were developed as an in vitro model for the acquisition of oestrogen and antioestrogen resistance. In this model there was no evidence that changes to ER receptor function and/or structure at the level of the ER gene, mRNA, ligand binding, and ability to induce progesterone receptor might account for the development of hormone resistance. However, the ability of ER to interact with a DNA sequence containing the vitellogenin promoter oestrogen response element, as assessed by gel retardation assay, was impaired in the clone showing the greatest degree of oestrogen and antioestrogen resistance.     

Alpha-fetoprotein and trisomy 21

In 31 affected pregnancies with Down syndrome, the median maternal serum alpha-fetoprotein value was lower than normal, 0.76 MoM, and median amniotic fluid value was quite normal, 0.98 MoM. Selecting an arbitrary cutoff-point of 0.5 MoM, 4.1 percent of normal gestations show values less than 0.5 MoM. Authors discuss problems about screening for fetal Down's syndrome by measuring maternal serum AFP levels.     

Purification and characterization of cytoplasmic creatine kinase isozymes ofXenopus laevis

The soluble creatine kinase isozymes CK-II, CK-III, and CK-IV fromXenopus laevis have been purified to apparent homogeneity and their subunits characterized by means of molecular weight, peptide pattern, and dissociation-reassociation experiments. CK-III and CK-IV are homodimeric isozymes whose subunits are distinct in both molecular weight (42,000 and 41,000, respectively) andStaphylococcus aureus V₈ peptide pattern. In dissociation-reassociation experiments, those two subunits do form active heterodimeric isozymes with one another or with rabbit M-CK subunits. Hybrid CK-III/IV isozymes occur also during embryonic differentiation and in adult heart muscle, whereas most other adult tissues contain only homodimeric CK-III or CK-IV isozymes. The CK-II isozyme is a heterodimer composed of one CK-III subunit and another subunit specific to CK-II (M _r =41,000). Neitherin vivo norin vitro does this subunit seem able to form homodimers or heterodimers with CK-IV and rabbit M-CK subunits. If we take into account the apparent association of CK-I isozyme with cellular organelles, these results corroborate earlier statements and suggest that the CK isozyme system ofX. laevis is encoded by at least four differentially regulated genomic loci.     

Vernonia galamensis,Potential new crop source of epoxy acid

Vernonia galamensis is a good source of seed oil rich in epoxy acid, which can be used to manufacture plastic formulations, protective coatings, and other products. Seed from a natural stand in Ethiopia contained 31% epoxy acid. Under cultivation in Kenya, this unimproved germplasm produced a substantial yield of seed with 32% epoxy acid. This African species has good natural seed retention and is a promising new crop for semiarid tropical areas.     

Genealogy of Principal Strains of the Yeast Genetic Stock Center

We have constructed a genealogy of strain S288C, from which many of the mutant and segregant strains currently used in studies on the genetics and molecular biology of Saccharomyces cerevisiae have been derived. We have determined that its six progenitor strains were EM93, EM126, NRRL YB-210 and the three baking strains Yeast Foam, FLD and LK. We have estimated that approximately 88% of the gene pool of S288C is contributed by strain EM93. The principal ancestral genotypes were those of segregant strains EM93-1C and EM93-3B, initially distributed by C. C. Lindegren to several laboratories. We have analyzed an isolate of a lyophilized culture of strain EM93 and determined its genotype as MATa/MATα SUC2/SUC2 GAL2/gal2 MAL/MAL mel/mel CUP1/cup1 FLO1/flo1. Strain EM93 is therefore the probable origin of genes SUC2, gal2, CUP1 and flo1 of S288C. We give details of the current availability of several of the progenitor strains and propose that this genealogy should be of assistance in elucidating the origins of several types of genetic and molecular heterogeneities in Saccharomyces.     

Evaluation of Diverse Antisera, Conjugates, and Support Media for Detecting Bradyrhizobium japonicum by Indirect Enzyme-Linked Immunosorbent Assay

We evaluated three antisera and four enzyme conjugates for the detection of Bradyrhizobium japonicum by an indirect enzyme-linked immunosorbent assay in microtiter plates. Nitrocellulose membrane sheets were then evaluated as an alternative support medium by using some combinations. Partially purified immunoglobulin G (IgG) or unpurified antisera to strain USDA 110 raised in rabbits, goats, or sheep was reacted in microtiter plates with alkaline phosphatase conjugated to protein A, goat anti-rabbit (GAR), sheep anti-rabbit (SAR), or rabbit anti-goat (RAG) IgG. Cultures or nodules containing homologous rhizobia were detected with equal sensitivity when protein A, GAR, or SAR was reacted with 5 μg of protein IgG per ml or a 1:800 titer of antisera from rabbits, but not goats or sheep. RAG reacted with IgG or antisera from goats or sheep. The detection limit was 2 × 10⁵ rhizobia per well. Rhizobia were spotted on nitrocellulose sheets as an alternative support medium, followed by soaking in 5 μg of protein per ml as IgG and 1:4,000 dilutions of protein A or GAR conjugate. Rhizobia in serogroup 110 were detected with the dye combination Nitro Blue Tetrazolium-5-bromo-4-chloro-3-indolyl phosphate (NBT-BCIP), and rhizobia in serogroup 122 were detected with fast red-naphthol phosphate (FR-NP). At the conclusion of the 5-h assay, purple (NBT-BCIP) or red (FR-NP) spots were visible in positive reactions. The sensitivity of detection was about 1,000 rhizobial cells or 3 μg of nodules tissue.     

Production of High-Viscosity Whey-Glucose Broths by a Xanthomonas campestris Strain

Acclimation of a sandy soil to an air-natural gas mixture stimulated the biological oxidation of chloroform to carbon dioxide. Acetylene and methane inhibited chloroform oxidation. Chloroform oxidation continued up to 31 days in the absence of methane. Chloroform oxidation rates increased at chloroform concentrations up to 5 μg g of soil^-1.