Optimization of SCoT-PCR reaction system and screening primers for cassava

ObjectiveSCoT-PCR reaction system of cassava was optimized and SCoT primers were screened in order to provide technical support for applying SCoT molecular marker technique in cassava molecule breeding. MethodTaking cassava variety(Xinxuan 048) as materials,the orthogonal design method was used to optimize five factors including DNA template amount,Mg2+concentrations,dNTPs concentrations,primer concentration and Taq DNA polymerase amount. Then the optimized SCoT-PCR reaction system was used to screen SCoT primers for cassava. ResultResults showed that,the optimal SCoT-PCR reaction system(20 μL) was composed of 60 ng DNA template,0.30 μmol/L primer,1.5 U Taq DNA polymerase,1.50 mmol/L Mg2+ and 0.25 mmol/L dNTPs. Nineteen primers were selected from 36 SCoT primers based on optimal SCoT-PCR reaction system,which could amplify steadily a large number of clear,distinguishable and polymorphic bands. ConclusionThe optimal SCoT-PCR reaction system shows stable detection result and good reproducibility ,and 19 selected primers can be used for genetic diversity analysis of cassava.