Abstract: The variation of SPAD value with doubled haploid lines in Brassica napus L. was analyzed in different ecological environments, to reveal the genetic regularity of chloroplast content, innovate new germplasm resources with higher chlorophyll content and establish new approach and theory for high photosynthetic efficiency breeding. In this paper, 170 DH lines in B.napus L. were planted in winter rapeseed area (Shaanxi Dali) and in spring rapeseed area (Gansu Zhangye) for three consecutive years. The results showed that the chlorophyll content had the genetic stability and was easily subjected to environment influence in DH lines of different genetic background or different DH lines materials of the same parents, the performance trend of SPAD value was basically the same in the same ecological environment of different years, but had extremely significant difference in different ecological environments. According to analysis of SPAD value frequency distribution for DH lines parents and DH lines, there was a wide segregation range and continuous distribution of chlorophyll content in two ecological environments in three consecutive years, and normal or near-normal distribution with transgressive segregation. These results also indicated chlorophyll content of B.napus L. was a quantitative trait and genetically controlled by many pairs of genes.