Abstract: Construction of subtracted cDNA library of Malus xiaojinensis is the base of screening and cloning genes under Fe-deficiency stress. Using LD-PCR technique, cDNA was synthesized from total RNA of Malus xiaojinensis roots treated under Fe-deficiency stress(EDTA-Fe2＋，4μmol/L)which was used as tester and Fe-sufficiency(EDTA-Fe2＋，40μmol/L)as driver. A subtracted cDNA Library was constructed using suppression subtractive hybridization(SSH). After two hybridizations and two suppression PCRs, the amplified library contained 1200 positive clones. Plasmid inserts were PCR amplified and showed 300~800bp inserts. There is the same result with RsaⅠdigestion of plasmid