“Mutational Analysis Of Mitochondrial Trna Proline Gene In Myoclonal Epileptic Patients From North Waziristan”

Abstract

The term "epilepsy" refers to a group of neurological diseases and syndromes that are characterized by recurring, spontaneous, paroxysmal seizure activity. Juvenile myoclonic epilepsy is a comparatively benign form of idiopathic generalized epilepsy. Genetics appears to have a major role in the genesis of epilepsy. The mitochondrion is the only organelle in the cell that has its own DNA that codes for 13 proteins. The mt-DNA circular genome is 16154bp in length. Mt-DNA mutations cause multisystem mitochondrial disorders having a diverse set of clinical signs and symptoms. More than 20-heteroplasmic point mutations has been linked to Myoclonic epilepsy with ragged red fibers, demonstrating the genetic variability of mitochondrial disorders. In Mt- DNA the most common mutation (m.8344A>G) is observed in tRNA lys (MT-TK) gene, which accounts for 80% of Myoclonic epilepsy ragged red fibers cases. The current study was designed to identify the mutation in Proline (MT-TP) gene of mt-DNA. This cross-sectional study was conducted in North Waziristan. Non-probability convenient sampling technique was adopted. After taking informed consent questioner was filled and 3mL of blood sample was collected in EDTA tube. A total of 38 samples were collected from 10 families using sample size calculation formula n=p(1-p2) z2/d2. The samples size was calculated by using p(prevalence)=4% (0.04), d (margins of error) =5% (0.05), z; confidence level=95% (1.96) which provided a sample size of (n=38). Thermo Scientific Gene-JET Genomic DNA Purification Kit was used for mt-DNA extraction. DNA quantity was confirmed by nano drop and agarose gel electrophoresis. Specific primers were designed for 394 bp target of proline gene and the extracted DNA was used as a template for the PCR amplification. The specific amplified region of proline-gene was confirmed through gel electrophoreses. The amplified PCR product bands were carefully removed, purified and cleaned through gene clean process. The amplified product was sequenced in BGI Hongkong for nucleotide. Out of total 14 epilepsy sample population, 10 (71.4%) were males and 4 (28.6%) were females. In the current study, no mutation was found in the (MT-TP) Proline-tRNA gene ranged from 16008-15956 bp of studied specimens. The results of over study suggest that mutation may be found in other region of MT-TP gene or MT-TK gene. The current study also recommends conducting the research on a wider scale to rule out the other mutation in different genes of mitochondrial genome of epileptic families in Pakistani population.