Molecular Cloning and Expression Analysis of Genes Encoding Two Microsomal Oleate Desaturases (FAD2) from Safflower (Carthamus tinctorius L.)

The endoplasmic reticulum (ER)-associated oleate desaturase (FAD2) is the key enzyme responsible for the production of linoleic acid in plants. In safflower, a seed-type FAD2 gene (CtFAD2-1) has been isolated previously. In this study, two different cDNA sequences, designated CtFAD2-2 and CtFAD2-3, encoding two microsomal oleate desaturases have been isolated from safflower (Carthamus tinctorius L.) using a polymerase chain reaction (PCR) approach. Both deduced amino acid sequences showed the three histidine boxes characteristic of all membrane-bound desaturases, and possess a C-terminal ER retention signal. Phylogentic analysis shows that CtFAD2-2 is grouped with other house-keeping type FAD2 sequences from oil crops, while CtFAD2-3 does not belong to seed type or house-keeping type FAD2. Real-time PCR analysis showed that CtFAD2-2 gene is constitutively expressed in both vegetative tissues and developing seeds, with higher transcript levels in roots, stems and petioles. Transcript of CtFAD2-3 was strongly detected in developing seeds, showing low levels in vegetative tissues. In response to low-temperature stress, CtFAD2-2 and CtFAD2-3 in different tissues showed a different behavior (up- or down-regulated). The transcript of CtFAD2-2 was dramatically decreased in roots, stems and petioles under low temperature, while CtFAD2-3 showed a significant increase in these tissues. On the contrary, in leaves, the strongly enhanced expression of CtFAD2-2 was observed under low temperature, while the transcript of CtFAD2-3 showed a little decrease.