Establishment Of A Method For Sudden Cardiac Death Related Gene Resequencing

At present,the clinical detection method of sudden cardiac death is mainly to screen family history and personal history,as well as physical examination,electrocardiogram examination,echocardiography and other examinations,but there are some shortcomings,such as low detection rate and lack of pertinence.In recent years,studies on the correlation between sudden cardiac death and genes have made it possible to screen genes for sudden cardiac death.Therefore,it is particularly important to avoid the occurrence of sudden cardiac death through early gene screening.The purpose of this study was to develop an approach based on gene resequencing to determine the genetic defects in patients with sudden cardiac death related disease.Using Agilent SureSelect target gene capture technology,design specific probe of 88 genes with 1727 exons,DNA fragments in the target genome region were enriched by hybridization with genomic DNA library,and sequenced by Illumina MiniSeq sequencer.Mutations in the target genes were found by means of biological information analysis.Then based on American College of Medical Genetics and Genomics(ACMG)guidelines to define the rate of mutations,find out pathogenic and likely pathogenic mutations,and provide clinical interpretation report.Finally,the variants were further validated by Sanger sequencing.Specifically designed and synthesized target gene-specific capture probes in this study can effectively capture and enrich target fragments of genomic DNA.The sequencing depth was determined by gradient screening,and the optimal sequencing depth was about 200×.In this study,4 patients with arrhythmia and 8 family members in an atrial fibrillation family were selected as subjects..The gene resequencing technology established in this study was used to detect 5 pathogenic or likely pathogenic mutations in 12 subjects,respectively:c.79C>T heterozygous mutation in KCNE2,c.677C>T heterozygous mutation in SCN5A,c.72845delA heterozygous mutation in TTN,c.2541de1C heterozygous mutation in SCN5A,c.988G>T heterozygous mutation in TRPM4.The results were verified by the currently recognized Sanger method,and it was found that the results of Sanger verification were consistent with the results of gene resequencing technology.Therefore,this study successfully established a new method to detect sudden cardiac death related genetic diseases--gene resequencing technology,which is accurate and effective,and saves time and cost.This study provides a theoretical basis for the clinical application of gene resequencing in the detection of sudden cardiac death related gene mutations in the future,provides new ideas for the diagnosis,screening and prevention of diseases,and provides a new method for family screening of proband family members and relatives.Therefore,this study has important clinical significance in the diagnosis and prevention of sudden cardiac death.