Lactobacillus plantarum can synthesize nano selenium with low toxicity and high efficiency during its growth and reproduction. Common nutrient substrates have great interference on the transformation of Lactobacillus plantarum into nano selenium. The experiment aims to avoid the influence of substrate on nano selenium transformation by modified medium and pre-washing treatment. The results showed that glucose and lactose have reducibility， which can convert sodium selenite into nano-selenium； the modified MRS medium was suitable for the growth of Lactobacillus plantarum and also conducive to the synthesis of nano selenium. When the mass concentration of sodium selenite was 4 g/L， the selenium enrichment rate was higher. The synthesized nano selenium particles were uniform， stable and well dispersed； Lactobacillus plantarum washed and inoculated in phosphate buffer can promote the conversion of sodium selenite into nano-selenium， and nanoparticles can be seen in the cell, which can avoid matrix interference and lay the foundation for the research on the transformation of nano-selenium by Lactobacillus plantarum.