Structure of REC2, a recombinational repair gene of Ustilago maydis, and its function in homologous recombination between plasmid and chromosomal sequences.

Mutation in the REC2 gene of Ustilago maydis leads to defects in DNA repair, recombination, and meiosis. Analysis of the primary sequence of the Rec2 protein reveals a region with significant homology to bacterial RecA protein and to the yeast recombination proteins Dmc1, Rad51, and Rad57. This homologous region in the U. maydis Rec2 protein was found to be functionally sensitive to mutation, lending support to the hypothesis that Rec2 has a functional RecA-like domain essential for activity in recombination and repair. Homologous recombination between plasmid and chromosomal DNA sequences is reduced substantially in the rec2 mutant following transformation. The frequency can be restored to a level approaching, but not exceeding, that observed in the wild-type strain if transformation is performed with cells containing multiple copies of REC2.     

Genetic markers for Atlantic salmon (Salmo salar L.): single locus inheritance and joint segregation analyses of minisatellite (VNTR) DNA loci

Relatively few genetic markers are available for detailed studies of Atlantic salmon. The detection of 12 distinct minisatellite DNA loci in this species (by 10 Atlantic salmon and brown trout derived probes) and subsequent inheritance analyses in two half-sib families are reported here. Disomic Mendelian inheritance was confirmed at all loci. Only a single aberrant progeny genotype (at Ssa -A60) was observed among 138 progeny screened. None of the loci was sex-linked. The tight linkage association Str -A22/1 with Str -A22/2, previously reported for brown trout, was found to be conserved in the Atlantic salmon genome. An additional male-specific linkage group, Ssa -A34 with Str -A9/2, was also noted. These highly polymorphic loci should find widespread use as chromosomal, individual, familial and, probably, population markers.     

Vaccination and the population structure of antigenically diverse pathogens that exchange genetic material.

Populations of antigenically diverse pathogens undergoing genetic exchange may be categorized into strains on the basis of a set of principal protective antigens. The extent to which polyvalent vaccines based on these protective antigens can alter the population structure of the pathogen is determined by the degree of cross-protection between strains. In the case where there is no cross-protection, vaccinating against a particular strain will have no effect on the others. As cross-protection increases, the strains containing the antigenic variants included in the vaccine will be diminished in prevalence, and those that do not will increase in prevalence. The rise in prevalence of the latter will become more and more exaggerated as cross-protection increases. However, beyond a critical level of cross-protection, in the absence of vaccination, the steady state of the system is asymmetric in that a certain subset of strains (with non-overlapping repertoires of antigenic variants) will dominate over the others in terms of prevalence. Under these circumstances, a vaccine consisting of the most immunogenic combinations of antigenic variants can cause a dramatic increase in frequency of a subset of rare strains.     

Initial evaluation of the non-sulfhydryl-containing converting enzyme inhibitor MK-521 in hypertensive humans

MK-521 is a new orally active, nonsulfhydryl angiotensin-converting enzyme (ACE) inhibitor. Single doses of 2.5, 5.0, 10.0, and 20.0 mg were administered to nine hypertensive patients alternating with placebo. All doses of MK-521 caused profound suppression of ACE activity for more than 24 h and decreased standing diastolic blood pressure for more than 12 h without changes in pulse rate. Although there was no further reduction in blood pressure with doses above 5.0 mg, the duration of action was prolonged for more than 24 h with the higher doses. Serum MK-521 concentrations increased with dosage, and ACE was inhibited maximally at concentrations above 10 ng/ml. In this initial study, MK-521 was well tolerated and proved to be a potent and long-acting antihypertensive agent.     

Slide staining device for use during space flight.

A slide staining device is described that performs Gram and Wright stains during space flight. Reagents and liquid wastes are contained within a closed system.     

Preservation of RNA for in situ hybridization: Carnoy's versus formaldehyde fixation.

Tissues fixed with organic solvent fixatives such as Carnoy's solution are known to give poor and erratic results with in situ hybridization, whereas those fixed with paraformaldehyde produce more consistent results. To understand this difference and to improve the utility of Carnoy's-fixed tissue for in situ hybridization, we explored several parameters of RNA integrity and preservation. Carnoy's-fixed, paraffin-embedded livers and paraformaldehyde-fixed, paraffin-embedded livers of mice were compared for RNA extractability, degradation, and hybridizability. In addition, retention of RNA in tissue sections after sequential in situ hybridization treatments was compared. RNA was found to be easily extractable from Carnoy's-fixed liver and was well preserved, with only slight degradation of high molecular weight RNA. Conversely, only a small percentage of the RNA was extractable from paraformaldehyde-fixed liver unless the tissue was digested with protease. The extracted RNA was well preserved, without detectable degradation. Sections of tissue fixed in Carnoy's solution subjected to in situ hybridization retained only about 10% of their original RNA content and gave correspondingly weak in situ hybridization signals. Formaldehyde-fixed tissues retained much more of the RNA (about 45%) and produced strong in situ hybridization signals. Treatment of Carnoy's-fixed tissue sections with vaporous formaldehyde increased retention of RNA and provided in situ hybridization signals comparable with those of paraformaldehyde-fixed tissues.     

Determination of 35S-aminoacyl-transfer ribonucleic acid specific radioactivity in small tissue samples

Rate determination of protein synthesis utilizing tracer amino acid incorporation requires accurate assessment of the specific radioactivity of the labeled precursor aminoacyl-tRNA pool. Previously published methods presumably useful for the measurement of any aminoacyl-tRNA were unsuccessful when applied to [³⁵S]methionine, due to the unique chemical properties of this amino acid. Herein we describe modifications of these methods necessary for the measurement of ³⁵S-aminoacyl-tRNA specific radioactivity from small tissue samples incubated in the presence of [³⁵S]methionine. The use of [³⁵S]methionine of high specific radioactivity enables analysis of the methionyl-tRNA from less than 100 mg of tissue. Conditions for optimal recovery of ³⁵S-labeled dansyl-amino acid derivatives are presented and possible applications of this method are discussed.     

Differential biosynthesis of polyunsaturated fatty acids by Tetrahymena supplemented with ergosterol.

Tetrahymena grown with foreign sterols such as ergosterol incorporate them into cellular membranes at the expense of the native compound, tetrahymanol. It is shown that cells grown with ergosterol have a lessened capacity to produce the polyunsaturated linoleic and gamma-linolenic acids from [14C]oleic acid. However, the same cells have normal capacities to introduce double bonds at C-6 into linoleate, alpha-linolenate, or cis-vaccenate. Thus, a presumed 12-desaturase is inhibited in the presence of ergosterol, while desaturation at C-6 is unaffected.