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

ABSTRACT

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.