The critical chilling temperature of cold-sensitive lima bean (Phaseolus lunatus L.) seedlings was determined to be approx. 8°C. The full-length cDNAs of PlSAD, PlFAD2, and PlFAD3 encoding three fatty acid desaturases were isolated from lima bean leaves. The PlSAD open reading frame (ORF) had 1,317 bp and a single intron of 601 bp, encoding a polypeptide of 438 amino acids that showing 86% homology with the plastidial stearoyl-acyl carrier protein desaturase in soybean. The PlFAD2 ORF contained 1,059 bp and was uninterrupted, encoding a polypeptide of 352 amino acids having 85% identity with the microsomal omega-6 desaturase GmFAD2-3 in soybean. PlFAD3 ORF had 1,116 bp and seven introns, encoding a polypeptide of 371 amino acids showing 91% homology with the microsomal omega-3 desaturase FAD3 in cowpea. PlSAD and PlFAD2 were expressed highly in leaves but poorly in roots and stems, while PlFAD3 was expressed in three tissues. All three genes were significantly induced in leaves by drought. PlSAD and PlFAD3 in leaves were down-regulated by high temperature and salinity, whereas PlFAD2 was up-regulated by the two stresses. The expression patterns of the three genes in lima bean leaves under suboptimal temperature, 18°C, were different from those under 8°C. The down-regulation of PlSAD transcript at chilling temperature might be an important factor contributing to chilling susceptibility for lima bean plants.