Induction of hairy roots and characterization of peroxidase expression as a potential root growth marker in sesame.

ABSTRACT

Using hypocotyl and cotyledon of sesame seedlings, hairy root cultures were established and cDNA coding for a peroxidase was cloned from the roots. The frequency of sesame hairy root formation was higher in hypocotyl (33.4%) than cotyledon (9.3%). Applicable levels of kanamycin and hygromycin as a selectable marker were 100 microg/mL and 30 microg/mL, respectively. The peroxidase cDNA showed relatively high sequence identity with and similarity to plant class III peroxidase family. The cDNA encoded polypeptide was identified with the presence of three sequence features 1) the putative 4 disulfide bridges, 2) an ER-targeted signal sequence in the N-terminus, and 3) two triplets, NXS for glycosylation. A real-time RT-PCR exhibited an abrupt increase in the peroxidase transcription activity after 4-week cultures of the sesame hairy roots and its highest level in 6-week cultured hairy roots. In contrast, the growth pattern of sesame hairy roots showed a typical sigmoidal curve. The active hairy root growth began after 2-week culture and their stationary growth phase occurred after 5-week culture. These results suggested that the peroxidase expression patterns at its transcription level could be used a potential indicator signaling a message that there will be no longer active growth in hairy root cultures. The sesame peroxidase gene was differentially expressed in different tissues.