Author/Editor		Krhin, Blaž
Title		Določanje A46G polimorfizma v vodilnem zaporedju eksona 1 gena za citotoksični T limfocitni antigen 4
Translated title		Determination of A46G polymorphism in the leader sequence of exon 1 of cytotoxic T lymphocyte antigen 4
Type		monografija
Place		Ljubljana
Publisher		Fakulteta za farmacijo
Publication year		2004
Volume		str. 51
Language		slo
Abstract		Cytotoxic T lymphocyte antigen 4 (CTLA-4) acts as an important negative regulator of T cell activation in the immune response. Specific CTLA-4 gene polymorphisms play an important role in the pathogenesis of autoimmune disorders. Many studies of different populations shows an association of CTLA-4 gene polymorphisms with autoimmune thyroid disease, mentioning a recent study on slovene population which shows impact of allele G on thyroid antibody production. Aims of our study were: l. to implement and optimise the determination of A49G polymorphism in the exon 1 of the CTLA-4 gene; 2.a. to determine the difference in A and G allele distribution in patients with autoimmune thyroid disease and control subjects; 2.b. to determine the difference in AA, AG and GG genotype distribution in patients with autoimmune thyroid disease and in control subjects. Target DNA sequence was amplified by polymerase chain reaction (PCR). In the procedure of optimisation we defmed PCR mixture concentrations as follows: 200 ng/microL DNA, 16 mE/microL Taq polymerase, 0,5 microM specific primers, 200 microM dNTP and 2,5 mM MgC12. Amplification of target DNA was performed by 25 cycles of: denaturation at 94°C for 30 sec, primer annealing at 55°C for 30 sec, primer extension at 72°C for 30 sec. Described polymorphism was assesed by restriction fragment polymorphism analysis. Mixture of 1 U/microL BbvI and 5 microL PCR poduct was incubated at 37°C for 3 hours. All products were visualised by polyacrilamide gel electrophoresis. We did not find any significant difference (P<0.05) of A and G allele distribution between patients with autoimmune thyroid disease and control subjects. We did not find any significant difference (P<0.05) of AA, AG and GG genotype distribution between patients with autoimmune thyroid disease and control subjects. Although, the portion of GG genotype in group of patients with autoimmune thyroid disease is twice as much as in group of control subjects.
Descriptors		THYROIDITIS, AUTOIMMUNE T-LYMPHOCYTES, CYTOTOXIC ANTIGENS, CD4 EXONS ALLELES POLYMERASE CHAIN REACTION ELECTROPHORESIS, POLYACRYLAMIDE GEL