The invention relates to a method for preparing novel germplasm of S gene homozygote by radioactive mutagenesis of pollen, which belongs to the field of molecular genetics. Seven S1 gene homozygote plants, four S2 gene homozygote plants, three S3 gene homozygote plants and two S5 gene homozygote plants are screened and appraised together from self-bred progeny plants by radiating huanghua and fengshui pear flower pollen with Co60 gamma 40Gy mutagenesis radiation. The plants of S gene homozygote provide good test piece for future researches on ploidy culturing of pear, cell signal transductionwhich identifies pollen and pistil mutually, time and space expression characteristics of S gene and function verification of protein product thereof, guide the field pollination and have good theoretical research value and production application.