Glycerol-salt Matrix Assisted Stacking Of Nucleic Acids For Capillary Electrophoresis

This work reported a new stacking effect for nucleic acids subjected to capillary electrophoresis when mixtures of glycerol and inorganic salts were present in the sample (not in separation buffer). Different types of nucleic acid samples including RNA, double strain DNA (dsDNA), as well as single strain DNA (ssDNA), were tested for the stacking effects. Under high concentration of salts and glycerol, the RNA sample could be stacked as high as 30 times.In this report we also have discussed the reasons for staking.Ribogreen is a special fluorescence dye that can stain dsDNA, ssDNA as well as RNA. In our recent efforts to develop a universal buffer system for analyzing interactions between nucleic acids with proteins by capillary zone electrophoresis, we found that 1×TBE buffer with Ribogreen can be used for separating nucleic acids–protein complex and highly sensitive detection of nucleic acids by laser induced fluorescence with a 488 nm Argon laser (data not shown). Therefore, this buffer is used throughout experiments.The role of glycerol played for the stacking was explained based on its ability to change nucleic acid chains'conformation and its serve as a terminating ion during a pseudo-transit isotachophoresis (ITP) step. The glycerol-salt matrix will offer more tolerance to salts in sample matrix, higher detection sensitivity and higher resolution of nucleic acids analyzed by Capillary Electrophoresis.Thus this glycerol-salt matrix will not only make it possible to introduce biological samples into capillary for analyzing nucleic acids directly without sample cleanup, but also could find particular applications for analysing sequence of DNA by Capillary Electrophoresis.