Optimization and application of sugarcane SSR-PCR system based on CGE technology

ObjectiveSSR-PCR reaction system of sugarcane based on capillary gel electrophoresis(CGE) technology was optimized to provide technical support for research on genetic diversity and genetic relationship, molecular-assisted breeding and genetic map construction of sugarcane. MethodFour factors affecting SSR-PCR were optimized by orthogonal exprement based on CGE technology, including dNTPs concentration, primer concentration, Taq DNA polymerase amount and DNA template amount. Then the optimal SSR-PCR reaction system was confirmed and applied to genetic analysis of 8 sugarcane germplasm resources. ResultThe results showed that, the optimal reaction system(20 μL) was composed of 0.15 mmol/L dNTPs, 1.5 U Taq DNA polymerase, 0.50 μmol/L primers and 25 ng DNA templates. Using optimal reaction system and 8 pairs of SSR primers, a total of 101 fragments were amplified from 8 sugarcane germplasms of several coun-tries, 93 of which was polymorphic, with a polymorphism rate of 92.1%. The cluster analysis showed that, coefficient of genetic similarity among 8 germplasms ranged from 0.521 to 0.871. All tested materials could be clustered into two groups at similarity coefficient of 0.614, three foreign germplasms viz., PINDAR, CP72-1210, CP84-1198 and ROC20, GT69-156 were clustered into the same group, but ROC26, GT02-237 and GT97-69 were clustered into another group. Con-clusionThe established SSR-CGE system is suitable for genetic diversity analysis and genetic mapping of sugarcane due to its stable detection result and good reproducibility.