Molecular cloning and characterization of auxin-regulated genes from mungbean hypocotyls during adventitious root formation

Adventitious root formation (rooting) in 'Berken' mungbean [Vigna radiata (L.) Rwiclz.] cuttings is stimulated by indole-3-acetic acid (IAA), To understand the molecular events that occur during IAA-induced adventitious root initiation, a lambda gt11 cDNA library was made from mungbean hypocotyls treated with 500 mu M IAA for 3 hours and differentially screened, Two cDNAs MII-3 and MII-4 were isolated, Southern analysis revealed that both cDNAs are encoded by different genes, Expression studies showed different patterns for both genes. Both MII-3 and MII-4 were highly expressed in IAA treated hypocotyls, whereas MII-4 was also induced in IAA treated epicotyls. There was no expression of either MII-3 or MII-4 in control or IAA treated leaves, With increasing concentrations of IAA from 100 to 1000 mu M there was an increase in the average root number per cutting as well as a stimulation in MII-3 and MII-4. Both MII-3 and MII-4 showed a stimulation in expression 4 hours following treatment with 500 mu M IAA reaching a maximum from 4 to 8 hours followed by a decline thereafter, Basal expression of MII-3 was evident between 2 and 8 hours, whereas, a high degree of basal expression was found with MII-4 from 1 to 8 hours followed by a sharp decline. Cycloheximide (50 mu M) dramatically reduced rooting and MII-3 expression, whereas MII-4 was only slightly affected.