Sensitive allele-specific real-time PCR test for mutations in BRAF codon V600 in skin melanoma.

Mutations at BRAF codon V600 are used as predictive biomarkers for targeted therapy of skin melanoma. Here, a simple sensitive test to detect mutations of BRAF-V600 was developed using real-time PCR with allele-specific primers and TaqMan probes. Two versions of the test using sense and antisense allele-specific primers were designed and evaluated.

[Detection of nonsense and frame shift mutations in human BRCA1 gene using new plasmid vector pPhoA-frame].

The technique for the detection of frame shift and nonsense mutations in BRCA1 gene was suggested. The technique presumes the construction of recombinant plasmids where the tested DNA fragment placed in-frame with alkaline phosphatase gene of Escherichia coli (phoA). A plasmid pPhoA-frame was constructed for such analysis, the plasmid contains DNA fragment coding for alkaline phosphatase of E.

[Guest peptide insertions into the surface loops of alkaline phosphatase].

The paper presents the description of the experiments in line with the rational concept for the "safe" insertion of guest polypeptides into the alkaline phosphatase with the minimal influence of the inserts on the enzymatic activity of the protein. Several approaches are described in the paper for the surface loop length estimation and two loops were used as the sites for guest peptides introduction by gene engineering technique. The experiments clearly demonstrate that insertions of several peptides after Ala218 of alkaline phosphatase (the site was selected by loop length analysis) do not block the activity of the enzyme.

Large family with both parents affected by distinct BRCA1 mutations: implications for genetic testing.

Although the probability of both parents being affected by BRCA1 mutations is not negligible, such families have not been systematically described in the literature. Here we present a large breast-ovarian cancer family, where 3 sisters and 1 half-sister inherited maternal BRCA1 5382insC mutation while the remaining 2 sisters carried paternal BRCA1 1629delC allele. No BRCA1 homozygous mutations has been detected, that is consistent with the data on lethality of BRCA1 knockout mice.

Limitations on the recombinant plasmid selection by Lac(+)/Lac(-) colony phenotype detection.

Lac(+)/Lac(-) selection of recombinant plasmids based on the insertional inactivation of LacZalpha gene cannot differentiate recombinant clones in some cases. Several fragments of exon 11 of human brca1 gene were cloned in LacZalpha-containing plasmids so that frameshift appeared at the 5(')-end of the fragments tested but these fragments were in frame with the part of LacZalpha situated downstream of the polylinker. All plasmids except one caused blue colonies formation after being transformed in Escherichia coli LacZDeltaM15 cells in spite of the frameshift.