A novel PCR-based technique called amplification fragment length polymorphism (AFLP) was developed and applied to Heve germlasm studies. The technique is based on the selective PCR amplification of restriction fragments from digested genomic DNA. There are more than six thousand species of germplasm in our academy germplasm garden and some of them are elite germplasm demonstrated by production and previous studies. AFLP fingerprinting analysis was performed on 25 clones (15 of Wickham clones and 10 of Amazon wild germplasm which respectively possess phenotype with high-yield/low-yield, cold-tolerant/cold-sensitive, Oidium-resistant/Oidium-sensitive, Tapping panel dryness (TPD)/health. The gels were run on a ABI PRISM DNA sequencer (P.E.Corp.) and the electrophoresis results were analyzed by using GeneScan TM Analysis software. AFLP reaction conditions for Hevea were optimized. The bands amplified sized 50-600bp and the fragment profiles of different clones were distinctive. The comparison with previous studies by using other molecular marker systems was discussed. AFLP fingerprints were highly reproducible and had great potential as a tool for evaluating genetic diversity and clonal identification of Hevea brasiliensis.