Moreover, hearing loss was diagnosed. The external genitalia were normal. Further examinations
at the age of click here 3 and 5 months showed normal psychomotor and somatic development (Fig. 1a). DNA was isolated from peripheral blood leukocytes of the patient and his healthy parents. Exons 1 through 27 of TCOF1, including exon–intron borders, were amplified by PCR under optimal conditions, using specific primers. The PCR products were subjected to multitemperature single-stranded conformation polymorphism (MSSCP) analysis at 5 °C, 15 °C and 25 °C, using the DNA Pointer Mutation Detection System. The electrophoresis was followed by silver staining. The PCR products were purified on the DNA GelOut columns (A&A Biotechnology, Poland) followed by direct sequencing with the use of a BigDye ver.3.0 dye terminator cycle sequencing kit and specific primers. The dideoxy-terminated fragments were identified by capillary gel-electrophoresis based on the ABI 310 DNA Analysis System. The MSSCP analysis of the amplified fragments of exon 13 of the TCOF1 gene demonstrated changes
in the electrophoretic mobility in this patient, while the changes were not observed in the patient’s parents. selleck compound In order to confirm the results obtained in MSSCP analysis a direct sequence analysis was performed. Sequence analysis demonstrated a novel, heterozygotic c.1978delC mutation in exon 13 of TCOF1. In the case of the patient’s parents direct DNA sequencing showed normal sequences ( Fig. 1b). A majority of mutations responsible for Treacher Collins syndrome are localized in exons, mainly in the hot spots in exons 10, 13, 15, 16, 23 and 24 [9]. The most commonly occurring mutations of the TCOF1 gene include deletions, which cause a shift of the reading frame, formation of the termination codon and shortening
of the protein Thiamine-diphosphate kinase product. The next most common mutations of the TCOF1 gene are insertions, the longest insertion localized on exon 5 [14]. In the presented patient a novel, heterozygotic deletion c.1978delC was detected in the TCOF1 gene. This mutation was absent in the patient’s parents which probably indicates a de novo origin. Analysis of the novel c.1978delC deletion with the use of the OMIGA 2.0 system indicates that it causes a premature termination of translation at 677aa, which results in the formation of a protein product of the gene devoid of the nuclear localization signal. We believe that these findings will facilitate precise diagnosis of the patient and will extend our knowledge on the pathogenesis of TCS. Molecular diagnosis of TCS is essential in prenatal and postnatal screening, being of great importance for genetic counseling as well. BAM-K – study design, data collection and interpretation, acceptance of final manuscript version, literature search. RS – data collection, acceptance of final manuscript version. MMS – acceptance of final manuscript version. None declared. None declared.