[Clinical and molecular genetic investigation of Waardenburg syndrome type 1]. / Klinicheskoe i molekuliarno-geneticheskoe issledovanie sindroma Vaardenburga I tipa]

Hypoacusis is a common sensory defect in humans which creates problems in communication. Heredity is essential in etiology of hypoacusis and deafness. Genes PAX3 and MITF were studied in patients with Vaardenburg syndrome in 14 unrelated families. Five mutation defects in the gene PAX3 were found. This provided the final diagnosis of the syndrome in these families.

Two novel slavic point mutations in the low-density lipoprotein receptor gene in patients with familial hypercholesterolemia from St. Petersburg, Russia.

Using PCR-single-strand conformation polymorphism analysis, followed by sequencing of the abnormal samples, two novel point mutations in the 5' end of the fourth exon of the low-density lipoprotein receptor gene were found in two Russian families with familial hypercholesterolemia. These missense mutations consist of C127W and C139G transitions and result in a loss of one of three disulfide bonds in the fourth cysteine-rich repeat of the ligand-binding domain of the low-density lipoprotein receptor. Hypercholesterolemia segregated with the identified mutations.

Two novel low-density lipoprotein receptor gene mutations (E397X and 347delGCC) in St. Petersburg familial hypercholesterolemia.

Familial hypercholesterolemia (FH), a monogenic disease known to be caused by low-density lipoprotein receptor (LDLR) gene mutations, results in the development of premature atherosclerosis and coronary artery disease in affected individuals. The spectrum of LDLR gene mutations in Russia is poorly known. Using polymerase chain reaction (PCR)-single-strand conformational polymorphism (SSCP) analysis, followed by DNA sequencing, we have screened selected exons of the LDLR gene in 80 unrelated St. Petersburg FH patients for the presence of mutations. Two new LDLR gene mutations, 347delGCC and E397X, were characterized among individuals with familial hypercholesterolemia in St. Petersburg. The carriers of both mutations possessed highly elevated blood serum cholesterol. Cosegregation of E397X mutation and LDLR gene RFLP haplotypes with hyperlipidemia was demonstrated by family study. Both mutations seem to be specific to Slavic patients.

[Absence of DNA polymorphisms on the region of the APOV gene coding the proposed low density lipoprotein-binding domain of the ApoB-100 protein]. / Otsutstvie DNK-polimorfizmov na uchastke gena APOB, kodiruiushchem predpolagaemyi domen sviazyvaniia belka ApoB-100 s retseptorom lipoproteidov nizkoi plotnosti.

To reveal the molecular basis of hereditary diathesis to hypercholesterolemia, single-strand conformation polymorphism (SSCP) analysis and subsequent sequencing were used to study structural organization of the APOB gene for apolipoprotein B-100 (ApoB-100). The gene region encoding a putative low density lipoprotein (LDL) receptor binding domain was analyzed in patients with myocardial infarction (MI). Differences in structure of the gene between patients with MI and healthy people were not detected.

Nonisotopic identification of two point mutations in the CYP21 gene responsible for nonclassic 21-hydroxylase deficiency.

A simple nonradioactive method was developed for identification of the Pro-30-Leu and Val-281-Leu mutant alleles in the CYP21B gene. Not only does this approach improve mutation analysis for patients with the late onset form of 21-hydroxylase deficiency, but it also decreases problems with interference by the CYP21A pseudogene sequence.

[Amplification of DNA for 15-30 minutes in single-use pipette tips]. / Amplifikatsiia DNK za 15-30 minut v odnorazovykh nakonechnikakh dlia mikrodozatorov.

Amplification of 200-1000-bp DNA fragments was performed in 15-30 min using a rapid thermal cycler based on the commercial instrument TC-1000-1 (IRLEN, St. Petersburg, Russia). Plastic pipette tips were used as thin walled, high surface to volume-ratio tubes, to increase the rate of heating (cooling) of 20 microliters samples, which allowed the time required for DNA amplification to be considerably reduced (5-10 times).

A rapid and simple DNA fingerprinting method using RFLP and SSCP analysis of the hypervariable noncoding region of human mitochondrial DNA.

A simple method for identification of individuals and maternity testing has been developed. This technique includes PCR amplification of the 199-bp hypervariable portion of the noncoding region of human mtDNA, digestion with RsaI and subsequent SSCP analysis of restriction fragments with DNA silver staining. Using this approach we have analysed the DNA fingerprint patterns of the family members. The fingerprints of maternally related individuals appeared to be identical in three generations, while maternally unrelated members of the family showed differences in their fingerprints, either in SSCP or both RFLP and SSCP patterns. Sequencing data have confirmed the results obtained. Further DNA fingerprinting analysis of 19 unrelated mother-child pairs by means of the method described revealed complete identity of the fingerprint patterns within the pairs. The probability of a random fingerprint match for two maternally unrelated individuals was estimated as 8%.