Establishment of the PCR Amplification and Sequencing System of Hsa-miR-17 Gene Promoter Region / 现代检验医学杂志

Objective To establish the PCR amplification and sequencing system of Hsa-miR-17 gene promoter region.Methods To establish the PCR amplification and sequencing system of Hsa-miR-17 gene promoter region and evaluate its performance by analyzing the sequence characteristics,designing PCR and sequencing primers,extracting sample genome DNA from different human cell lines and using optimization strategy of adding DMSO to the final concentration of 5 ％ and making use of 5' tailed PCR primers;Then the established system was further verified in 10 other cell line samples.Results The PCR amplification and sequencing system of hsa-miR-17 gene promoter region,which had good repeatability,good specificity and its detection limit was 10 ng/μl,was established successfully under the condition of using 5' tailed PCR primers and 5 ％ DMSO and that was confirmed by electrophoresis analysis and Sanger sequencing.And fragment of Has-miR-17 gene promoter region in 10 human cell lines were successfully achieved and verified by this system.Conclusion The PCR system of Hsa-miR-17 gene promoter region was established successfully.

Mutation Analysis of EGFR,ALK and ROS1 in Tumor Tissues of Patients with Non-Small Cell Lung Cancer in South of China / 现代检验医学杂志

Objective To investigate the characteristics of EGFR,ALK and ROS1 mutations in patients with non-small cell lung cancer (NSCLC) in South of China and its relationship with clinical features.Methods The tumor tissues and corresponding clinical data of 76 NSCLC patients in South of China from November 2016 to June 2017 were collected.The mutations of EGFR,ALK and ROS1 were detected by ARMS assay with Joint detection kit.Meanwhile,the correlation between gene mutation rate and clinical features was analyzed.Results The mutation rate of EGFR was 67.3％ (42/76) in 76 patients with NSCLC in South of China,19 del and L858R mutations were the main mutation types.There was a co-mutation including 19 del and L858R.The positive rate of ALK gene fusion was 17.1％ (13/76),and 4 cases of ALK gene fusion combined with EGFR mutation were detected.The positive rate of ROS1 gene fusion was 1.3％ (1/76),and there was no co-mutation with other genes.Compared with ROS1,EGFR and ALK mutation rate was higher,the difference was statistically significant (x2 =54.515,P =0.000;x2 =11.329,P =0.001).The mutation rate of EGFR in non-smoking NSCLC patients was significantly higher than that in smokers (x2 =4.578,P=0.032),while the mutation rate of ALK and ROS1 was not statistically significant (x2=0.000,P＞0.05).There was no statistically significant difference in EGFR,ALK and ROS1 gene mutation rates among NSCLC patients of different age,sex and histology (x2 =0.000 ～ 2.219,P＞ 0.05).Conclusion EGFR,ALK and ROS1 gene mutations can be seen in patients with NSCLC in South China,in which EGFR and ALK gene mutation rate is higher.