Abstract: Using SSR-PCR molecular technique, the genetic structure of 22 selected lines from F6 and F7 generations from 5003 × A19 were analyzed. In this study, 48 out of 88 pairs of SSR primers were selected, which gave stable and polymorphic amplification profiles in focus. The UPGMA analysis classified 22 selected lines and their parentes into the same cluster-PA，which included standard-testers 478 and B73.The average genetic background of 22 selected lines had 64.03% from temperate zone maize 5003,30.03% from tropical zone maize A19,and 5.94% not from parentes.The result showed,some new alleles were tested in some lines.This is a availably approach of maize germplasm enhancement improvement.