Atom probe tomography (APT) and transmission electron microscopy (TEM) were employed simultaneously to study nanoparticles and their strengthening effect in artificial aging (AA) Al-Mg-Si alloy with different natural aging (NA). We discovered that NA leads to formation of different type of nanoparticles meanwhile affect their fraction and dimension. Separation of the contribution from different types of nanoparticles to the alloy strength is conducted. In LNA-AA, β′ with a volume fraction of 0.16% contributions 54MPa to the yield strength while 0.14% β″ contributes 101MPa, which fully demonstrates that β″ is the main strengthening phase. The thermodynamic analysis was conducted for further research and the differential scanning calorimeter (DSC) results show that LNA forms more numerous clusters Ⅰ than SNA and consumes more vacancies and solute atoms then does not have enough driving force to precipitate clusters Ⅱ which are the nucleation sites of nanoparticles. When there is a certain distance between the nucleation sites, preferential growth of few β″ occur then converted to β′. The solute atoms in the region nearly which there is no nucleation site do not have enough driving force to diffuse and precipitate but stay in the matrix, reducing the volume fraction of the nanoparticles.