The invention provides micro spherical silica gel of 20 to 100micron which is used for liquid chromatography and a preparation method thereof. By taking tetraethyl orthosilicate as the materials and taking solid grain of silica sol or silicon dioxide as seed, efficient and fixed phase which is used for the chromatography is prepared by reaction of seed growth method; product performance is overall enhanced by carrying out preference in selecting the seed and catalyst as well as pore-expanding agent. The porous microsphere prepared by the method is characterized in that: height evenness is provided that 85 to 90 percent of the microsphere have the diameter between 20 to 100 micron; the aperture is between 2 to 60 nanometers; the pore volume is between 0.4 to 1.2ml/g. The porous microsphereis suitable for being used as filling substrate of the high-efficiency liquid chromatography and can meet the needs of vector of high-efficiency catalyst and high-efficiency adsorbent. The characteristic in height evenness of the grain diameter makes the porous microsphere especially suitable for serving as the filling substrate of the high-efficiency liquid chromatography.