p. azarfam

Recent molecular studies on Iranian β-thalassemia genes revealed the presence of eight common mutations associated with thalassemia. Although these mutations are frequent, there are other rare and unknown mutations that can create large problems in designing preventive programs. We detected and explained the common mutations in north-western Iran previously and detection of the rare and unknown mutations could be useful in diagnosis and design of future preventive programs.

In this study, 5ml peripheral blood from 20 Azari- β-thalassemia patients whose mutation was not revealed in the previous study was collected and DNA extraction was done by isopropanol and proteinase k method. Initially, samples were examined for the rare mutations: Codon6, Codon16, Codon41/42, Codon36/37, -88 and Codon22 by ARMS – PCR techniques and then the unknown cases were directly sequenced.

According to our results, Codon15(TGG-TGA), Codon16(-C), Codon36/37(-T), IVSII-848(C-A), IVSII-745(C-G), -28(A-C( and Codon25/26(+T) were recognized and added to the spectrom of beta globin gene mutations in Azerbaijan and Iran. Also, we detected four SNP sites: 5’UTR+20(C-T), Codon2 (CAC-CAT) , IVSII-16(C-G) and IVSII-666(T-C) in β-thalassemia genes.

Our results could be useful for developing molecular screening plans and help prenatal diagnosis of beta thalassemia in Azerbaijan , Iran and other neighboring countries.

β –Thalassaemia was first explained by Thomas Cooly as Cooly’s anaemia in 1925. The β- thalassaemias are hereditary autosomal disorders with decreased or absent β-globin chain synthesis. The most common genetic defects in β-thalassaemias are caused by point mutations, micro deletions or insertions within the β-globin gene.

In this research , 142 blood samples (64 from childrens hospital of Tabriz , 15 samples from Shahid Gazi hospital of Tabriz , 18 from Urumia and 45 samples from Aliasghar hospital of Ardebil) were taken from thalassaemic patients (who were previously diagnosed).Then 117 non-familial samples were selected . The DNA of the lymphocytes of blood samples was extracted by boiling and Proteinase K- SDS procedure, and mutations were detected by ARMS-PCR methods.

From the results obtained, eleven most common mutations,most of which were Mediterranean mutations were detected as follows IVS-I-110(G-A), IVS-I-1(G-A) ،IVS-I-5(G-C) ,Frameshift Codon 44 (-C,(codon5(-CT),IVS-1-6(T-C), IVS-I-25(-25bp del) ,Frameshift 8.9 (+G) ,IVS-II-1(G-A) ,Codon 39(C-T), Codon 30(G-C) the mutations of the samples were defined. The results showed that Frameshift 8.9 (+G), IVS-I-110 (G-A) ,IVS-II-I(G-A), IVS-I-5(G-C), IVS-I-1(G-A) , Frameshift Codon 44(-C) , codon5(-CT) , IVS-1-6(T-C) , IVS-I-25(-25bp del) with a frequency of 29.9%, 25.47%,17.83%, 7.00%, 6.36% , 6.63% , 3.8% , 2.5% , 0.63% represented the most common mutations in North - west Iran. No mutations in Codon 39(C-T) and Codon 30(G-C) were detected.

Cunclusion: 

The frequency of the same mutations in patients from North - West of Iran seems to be different as compared to other regions like Turkey, Pakistan, Lebanon and Fars province of Iran. The pattern of mutations in this region is more or less the same as in the Mediterranean region, but different from South west Asia and East Asia.