Proliferation and rol gene expression in hairy root lines of Egyptian clover

Abstract

Hairy roots were efficiently induced from seedlings of Egyptian clover (berseem clover, Trifolium alexandrinum L.) by infection with Agrobacterium rhizogenes strain DC-AR2 (harboring mikimopine-type pRi1724) and cultured on Murashige-Skoog's (MS) medium containing 1 mg l−1 naphthaleneacetic acid (NAA). Each of the hairy root lines showed variations in growth on solid MS medium containing various concentrations of NAA and these variations were also found in culture in liquid MS medium containing 0.5 mg l−1 NAA. To investigate whether the growth capacity is related to the endogenous auxin IAA and the expression of oncogenes on Ri plasmid T-DNA, the concentration of IAA was determined by ELISA using IAA-specific antibody and the extent of transcripts of oncogenes rolB and rolC was analyzed by Northern blot hybridization. From these results, in hairy roots of Egyptian clover, a rolB expression seemed to influence the intrinsic growth capacity and to call for the proper level in order that the hairy roots grew vigorously, while the endogenous IAA seemed to directly affect the growth capacity on medium without exogenous auxin.