Sumo4 Gene 163a / G Polymorphism And Chronic Complications Of Diabetes-related Research

Part 1 A Study of Conditions Influencing Polymerase Chain Reaction-Restriction Fragment Length Polymorphism in the Site of Small Ubiquitin-Like Modifier 4 Gene 163 A/G PolymorphismObjective To determine the optimum experimental conditions of 163 A/G polymorphism in SUMO4 gene detected by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP).Methods The main influential factors were detected by PCR in serial concentration gradient and electrophoresis,the main factors were detected by RFLP in different systems.Results(1) When denaturation temperature was 94℃could achieved the clearest electropherogram.(2) When annealing temperature was 55℃,PCR product level was higher than others and less non-specific products.(3) When primer concentration was 0.25umol/L,PCR product level was as well as others but had less dimer of primers.(4) PCR product level was highest when magnesium concentration was 1.5mmol/L.(5) When concentration of dNTP was 0.25mmol/L could got the most PCR product.(6) Taq enzyme was 2.5U could achieved a clear electropherogram and saved experimental materials.(7) It was 10U Mse I that could digest 10μl PCR product in 31μl digestion system.Conclution The optimum conditions:denaturation temperature was 94℃; annealing temperature was 55℃;primer concentration was 0.25umol/L;magnesium concentration was 1.5mmol/L;concentration of dNTP was 0.25mmol/L;Taq enzyme was 2.5U;the effective system was 31μl including 10μl DNA solution and 10 U enzyme. Part 2 A study on the association of SUMO4 gene 163 A/G polymorphism with diabetic nephropathyObjective To investigate the association of 163 A/G polymorphism in SUMO4 gene with type 2 diabetes and diabetic nephropathy.Methods The polymorphism of SUMO4 gene 163 A/G was detected by PCR-RFLP method in 232 unrelated patients with type 2 diabetes,including 103 normal albuminuria(DN0 group),65 microalbuminuria(DN1 group),64 macroalbuminuria(DN2 group) and 102 normal controls(NC group).The genotype frequencies,allele frequencies and clinic indexes were compared among groups.Results Except the GA genetype in DN group (DN1+DN2) was higher than those in DN0(χ~2=4.315,P=0.038),there was no differerce between other genetype and allele frequencies.Conclusion The GA genetype of SUMO4 gene may associate with diabetic nephropathy. Part 3 A study on the association of SUMO4 gene 163 A/G polymorphism with diabetic retinopathy and diabetic peripheral neuropathyObjective To investigate the association of 163 A/G polymorphism in SUMO4 gene with diabetic retinopathy and diabetic peripheral neuropathy.Methods The polymorphism of SUMO4 gene 163 A/G was detected by PCR-RFLP in 193 unrelated patients with type 2 diabetes and 102 normal controls.The genotype frequencies,allele frequencies and clinic indexs were compared among groups.Results There was no differerce between genetype and allele frequencies.Conclusion The SUMO4 gene 163 A/G polymorphism may not associate with diabetic retinopathy and diabetic peripheral neuropathy.