PCR-based detection of genetically modified soybean and maize in raw and highly processed foodstuffs.

The PCR method has proved to be an invaluable tool for the specific and sensitive detection of genetically modified material (e.g., Roundup Ready Soybean and Bt-176 "Maximizer" Maize) in foodstuffs. The first step in the procedure, namely the purification of nucleic acids from the sample, is often the deciding factor in the production of meaningful results. In this study, we present two procedures that enable an efficient isolation of trace amounts of genetic material from both raw and highly processed foodstuffs. We show that for optimal, PCR-ready DNA purification from highly processed foodstuffs and PCR inhibitor-rich substances--such as cocoa-containing products--adapted protocols for the QIAGEN QIAamp DNA Stool Mini Kit can be utilized. For complete DNA isolation from raw foodstuffs, a protocol using the DNeasy Plant Mini Kit is presented.

[Progression of intestinal secretory immunoglobulin A and the condition of the patients during naturopathic therapy and fasting therapy]. / Verläufe des sekretorischen Immunglobulins A des Darms und Befindlichkeit von Patienten unter naturheilkundlicher Therapie und Heilfasten.

BACKGROUND: In clinical observations it was noticed that patients who have been treated with naturopathic methods - especially with the treatment of heal fasting - experience a stabilization of their immune system. The immune system stands in close relation to the intestinal flora and influences the condition of the patient. QUESTION: For this reason the influence of a 3-week in-hospital naturopathic therapy - among others a heal-fasting therapy -, a corresponding follow-up on the intestine-associated immune system as well as the condition of the patients have been examined. PATIENTS AND METHODS: Classical naturopathic treatments based on standardized therapy concepts were applied. The patients (n = 55) showed different kinds of basic diseases, whereof in 56% of the cases the patients suffered of diseases of the skeleton, the muscles, and the connective tissue. The patients got a 3-week standardized high-quality nutrition or they took part in a juice-modified fasting therapy. The state of the local endogenous defense system within the intestine was measured by the concentration of the secretory immunoglobulin A (sIgA) in the feces of the patients. A positive criterion was a sIgA concentration in the stool of under 0.5 mg/g. Additionally the judgement of the patients concerning their own condition ('quality of life') was determined by the condition scale. RESULTS: It was shown that the content of the sIgA within the stool was increased during the course of the in-hospital stay, in the complete population of patients as well as in the different subgroups 'with heal-fasting' and 'without heal-fasting'. Even in the follow-up analysis it could be observed that 3 months after the treatment the content of secretory IgA in the stool of all groups was between 0.78 and 0.89 mg/g. This value was significantly higher than the starting values. The most intensive effects concerning the change of sIgA were measured within the group of the patients who performed heal fasting (epsilon > 1.0). Aside from the increase of the secretory IgA, a mediocre improvement of the condition of the patients within the pre-post comparison could be noticed, but no correlation between the secretory IgA values and the values concerning the condition was found. CONCLUSION: A 3-week in-hospital therapy with classical naturopathic treatments (but especially the application of the heal-fasting therapy) leads to a significant improvement of the immune condition of the intestine and consequently of the whole organism. The improving effect lasts beyond the stay in hospital.

Facilitated isolation of rare recombinants by ligase chain reaction: selection for intragenic crossover events in the Drosophila optomotor-blind gene.

Ligase chain reaction (LCR) was evaluated as a tool for the detection of point mutations. For the mutation studied, the specificity of the method is sufficient to detect the mutant allele in the presence of a 200-fold molar excess of the wild-type sequence. LCR was therefore employed in a genetic recombination experiment as a probe for a recessive lethal point mutation. LCR greatly facilitated the isolation of a rare recombinant originating from a crossover event in the 40 kb interval separating the lethal mutation and an enhancer trap insertion in the optomotor-blind locus. The recombinant will allow the study of gene control in situ, in a largely unperturbed regulatory environment.

Transcript identification in the optomotor-blind locus of Drosophila melanogaster by intragenic recombination mapping and PCR-aided sequence analysis of lethal point mutations.

The optomotor-blind gene of Drosophila melanogaster is large and genetically complex. Five partly independent complementation groups are uncovered by several viable and lethal mutations at the locus. At least 15 RNA signals have been detected by Northern blot analysis. One of them, T3, derived from a 75 kb primary transcript, has been proposed as the carrier of optomotor-blind function, based on the large size of its precursor and its tissue distribution. We here provide direct evidence that T3 is the optomotor-blind transcript. A facile and generally applicable selection scheme for the isolation of intragenic meiotic recombinants was applied to map two lethal optomotor-blind point mutations to exons of the T3 transcript. Amplification of mutant DNA by the polymerase chain reaction (PCR) and sequencing of the amplified exons revealed the presence of mutations that lead to truncation of the T3 open reading frame. The recombination rate observed in the optomoter-blind locus is within the range of rates that have been determined in a few other Drosophila loci.