Cystic fibrosis typing with DNA probes and screening for ΔF508 deletion in families from Southern France

Summary A sample of 235 individuals from 49 French cystic fibrosis (CF) families with at least one living affected child was typed with probes for restriction fragment length polymorphisms (RFLPs) known to be linked to the CF gene, and was screened for the ΔF508 mutation. Using a combination of six probes, 44 out of the 49 families were sufficiently informative to enable prenatal diagnosis or carrier determination. As in many other populations, linkage disequilibrium was found between the CF locus and the haplotype B (XV2c: allele 1; KM19: allele 2), which accounts for about 78% of CF chromosomes in our families. The ΔF508 deletion was present in 64.3% of CF chromosomes.     

Somatic hybrid plants produced by electrofusion between dihaploid potatoes: BF15 (H1), Aminca (H6) and Cardinal (H3)

In order to regenerate somatic hybrids, mesophyll protoplasts from a dihaploid potato, BF15 (H1), were electrofused with those from two other dihaploid clones, Aminca (H6) and Cardinal (H3). Determination of the ploidy level by flow cytometry showed that 10% of plants regenerated from the fusion experiment with BF15 + Aminca were diploids, 14% triploids, 63% tetraploids and very few were mixoploids or had a higher ploidy level. Using morphological markers and vigour in plant growth, we were able to recover a total of 24 somatic hybrid plants, respectively 20 and 4 hybrids (accounting for 12% and 13% of regenerants) from the fusions BF15 + Aminca and BF15 + Cardinal. Most of the somatic hybrids were at the expected tetraploid level (2n=4x=48). The hybrid nature was confirmed by examining isoenzyme patterns for malate dehydrogenase (MDH) and isocitrate dehydrogenase (ICD).     

Oligonucleotides covalently linked to intercalating agents. Influence of positively charged substituents on binding to complementary sequences

A pentamethylene chain was used to covalently link the 3'-phosphate of oligothymidylates to the 9-amino group of an acridine derivative. Positively charged substituents were further attached to the 3'-phosphate group to form 3'-phosphotriesters. These molecules form specific complexes with poly(rA) which involve the formation of a number of A X T base pairs equal to that of thymines in the oligonucleotide. Absorption changes induced in the acridine absorption bands are similar to those expected upon intercalation of the acridine dye between A X T base pairs. The acridine covalently linked to the 3'-phosphate strongly stabilizes the complexes formed with poly(rA) as compared with the corresponding unsubstituted oligodeoxynucleotide. The presence of a positively charged substituent on the 3'-phosphate together with the acridine dye further enhances the interaction. The effect of salt concentration on complex stability depends on the number of negatively charged phosphate groups of the oligodeoxynucleotide and on the nature of the substituents borne by the 3'-phosphate group. When the oligothymidylate is substituted by an acridine dye, the stability of the poly(rA) complexes increases when salt concentration increases. If an additional positively charged substituent is present on the 3'-phosphate group, stability decreases when salt concentration increases for the shortest oligonucleotide (trimer) and increases with longer oligonucleotides. Thermodynamic parameters have been calculated from the concentration dependence of melting temperatures.     

Spin probe clustering in human erythrocyte ghosts

Summary A model has been developed for 5-nitroxide stearate, I(12,3), distribution in human erythrocyte ghosts which accurately predicts ESR spectral alterations observed with increased probe/total lipid (P/L) at 37°C. This spin probe occupies a class of high-affinity, noninteracting sites at low loading. Saturation occurs with increasing probe concentration, and, at higher loading, the probe inserts itself at initially dilute sites to form membranebound clusters of variable size. No low probe remains at high P/L where all I(12,3) clusters in a concentrated phase. This model allows determination of the dilute/clustered probe ratio, and shows that I(12,3) segregates in erythrocytes at what might otherwise be considered low P/L (e.g., 1/359). These findings validate the earlier use of empirical parameters to estimate probe sequestration in biological membranes.     

Heterogeneous production of laccase by mycelium of white-rot fungi

Mycelium of white-rot fungi secretes laccase into the medium. It was found by cultivation on malt-agar plates that the mycelium does not produce laccase equally in all its parts. The youngest hyphae at the margins of the colony represent usually the maximum producers, whereas older hyphae produce less or none at all. An exception here isCollybia velutipes which is the weakest producer of laccase of all the fungi studied and where only the older hyphae begin to secrete it. Manometric estimation of laccase showed that maximum specific activity of laccase is achieved at the boundary between the phases of initial and linear growth and i₁₁ some cases during the first half of linear growth. Ageing of the mycelium characterized by certain changes in its metabolism is reflected in changes of enzyme production by fungal hypha of different age.     

Diet and urinary estrogen profile in premenopausal omnivorous and vegetarian women and in premenopausal women with breast cancer

The urinary estrogen profile was studied in the midfollicular phase twice, and diet four times during 1 yr in 10 premenopausal breast cancer (BC) patients consuming an omnivorous normal Finnish diet and in two control groups, one consuming an omnivorous (n = 12) and the other a lactovegetarian (n = 11) diet. Total fat intake in relation to caloric intake was almost identical in all three groups. Only with regard to grain fiber intake did the BC patients differ significantly from both other groups. No differences were found between the groups with regard to urinary excretion of 13 individual estrogens and total estrogens, with the exception of 4-hydroxyestrone (4-OH-E1), which was significantly lower (P less than 0.05) in the BC group than in the vegetarians. A high carbohydrate to protein ratio in the diet had a negative correlation with the excretion of 2-hydroxyestrogens and 2-hydroxyesterone (2-OH-E1) to 4-OH-E1 ratio. The BC group had significantly higher urinary 2-OH-E1 to E1 ratio (P less than 0.05) compared to the vegetarians. The 2-OH-E1 to 4-OH-E1 ratio was highest in the BC group (= 7.1) and differed significantly from that of the omnivores (= 4.3; P less than 0.02) and vegetarians (= 3.6; P less than 0.005). This ratio showed a negative correlation with intake of carbohydrates, starch, total and grain fiber. Urinary excretion of 4-OH-E1 correlated positively with total and grain fiber intake and plasma SHBG. Protein intake correlated positively with urinary 2-methoxy-E1 excretion, and retinol intake positively with catechol estrogen, E1 and E2 excretion. It is concluded that estrogen production and urinary estrogen profile in premenopausal breast cancer patients is normal with the exception of a low 4-OH-E1 excretion and high urinary 2-OH-E1 to 4-OH-E1 ratio. This ratio, which seems to depend on diet, is the only urinary estrogen parameter separating premenopausal BC patients from the control omnivorous and lactovegetarian women.