Human initiation protein Orc4 prefers triple stranded DNA.

In higher eukaryotes mechanism of DNA replication origin recognition and binding by origin recognition complex (ORC) is still unknown. Origin transfer studies have shown that origin sites are genetically determined, containing functionally interchangeable modules. One of such modules from the human lamin B2 origin of replication has the ability to adopt unorthodox structure partly composed of intramolecular triplex. Sequences involved in triplex formation coincide with ORC binding sites both in vitro and in vivo. To explore potential significance of unorthodox DNA structures in origin recognition by ORC, we tested DNA binding properties of human ORC subunit 4 (HsOrc4) which has independent DNA binding activity in vitro and similar binding characteristics as ORC holocomplex. Our results demonstrated that DNA binding activity of HsOrc4 depends on length and structure of DNA with triplex being the protein's preferred binding target. Such feature could play part in origin selection through directing ORC to DNA sequence prone to adopt unorthodox structure.

Parental reports of emotional and behavioural difficulties on the SDQ for school-age children with vertically acquired HIV infection living in London.

This study uses a screening questionnaire (the Strengths and Difficulties Questionnaire, SDQ) to indicate levels of parental or carer concern about behavioural or emotional difficulties in a representative group of school-age children with vertically acquired HIV attending London clinics. Fourteen per cent of the total 107 children screened were reported to have behavioural and emotional difficulties scoring in the "abnormal" range. Older children tended to have higher scores. Overall, levels of reported difficulties were found to be similar to those reported for other chronic childhood illnesses and slightly higher than in the general child population.

Analysis of common CFTR polymorphisms 5T, M470V and R75Q in healthy Serbian population.

Three common CFTR polymorphisms, 5T, M470V and R75Q, have been shown to be relatively frequent in Serbian patients with monosymptomatic CF disorders. Since there is a variation in distribution of common polymorphisms among different populations, it was important to compare their frequencies in patients with the frequencies in healthy population in order to assess the possible role of these polymorphisms in the monosymptomatic CF disorders. Samples obtained from 100 healthy Serbian individuals were analyzed for the presence of CFTR 5T, M470V and R75Q variants by PSM, RFLP and DGGE methods, respectively. Allele 5T was present in two individuals, giving the allelic frequency of 1% (2/200 alleles). The frequency obtained for allele M470 was 45% (90/200 alleles), while V470 allele was present with the frequency of 55% (110/200 alleles). Polymorphism R75Q was present in two individuals, with allelic frequency of 1% (2/200 alleles). Our study has shown that the frequencies of two common polymorphisms, 5T and M470V, differ significantly in Serbian population in comparison with other South European populations. Since it appears that Serbian population has a specific distribution of studied CFTR gene variants, it would also be interesting to analyze other common variants of this gene in our population. Such data can also be potentially useful as anthropogenetic markers in population studies.

PCR amplification on whole blood samples treated with different commonly used anticoagulants.

The aim of this study was to explore the influence of different anticoagulants on PCR amplification, as well as to optimize PCR amplification of blood samples for successful mutation detection. For the purpose of this study the amplification of part of methylenetetrahydrofolate reductase (MTHFR) gene exon 4, in which mutation C677T is located, was performed. With the exception of sodium heparin, the presence of other commonly used anticoagulants (sodium citrate, K3EDTA, lithium heparin) made it possible to obtain satisfactory amplification. The described method, apart from saving time and material, enables successful PCR-based analysis even when a very small amount of blood sample is available.