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| Study on the experimental conditions for DDAH2 gene rs805304 polymorphism detection with PCR-RFLP |
| SHI Rou1 DONG Rongjing1 LIU Hua2 LI Huifang1▲ |
1.Deparment of Diabetes, the First Affiliated Hospital of Kunming Medical University, Yunnan Province, Kunming 650032, China;
2.Department of Clinical Laboratory, the First Affiliated Hospital of Kunming Medical University, Yunnan Province, Kunming 650032, China |
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Abstract Objective To investigate the optimum experimental conditions for dimethylarginine dimethylamine hydrolase 2 (DDAH2) gene rs805304 polymorphism by polymerase chain reaction - restricted fragment length polymorphism (PCR-RFLP). Methods The DDAH2-rs805304 locus polymorphism in 182 T2DM cases admitted to the First Affiliated Hospital of Kunming Medical University from January 2016 to March 2017 and 147 healthy controls were detected by PCR-RFLP. The main factors influencing PCR-RFLP were studied. Results The optimum conditions: annealing temperature was 58℃, denaturation temperature was 95℃, 2× Power Taq PCR Master Mix concentration was 12.5 μL, primer concentration was 0.8 μmo/L, the effective system was 15 μL including 8 μL DNA solution and 5 U restriction enzyme BstUI. Conclusion The exploration of optimal experimental conditions has laid a foundation for the subsequent research.
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