MORPHOLOGICAL STRUCTURED MODEL FOR HAIRY ROOT CULTURES I. BERZINl, D. MILLS2, J.e. MERCHUK3 I. Department of Chemical Engineering, Ben-Gurion University of the Negev, Beer-Sheva, Israel. 2. Institutes for Applied Biology, Ben-Gurion University of the Negev, Beer-Sheva, Israel. 3. Unit of Biotechnology, Ben-Gurion University of the Negev, Beer-Sheva, Israel. INTRODUCTION The possibility of using hairy root (HR) cultures for producing secondary metabolites on a large scale has recently received a great deal of attention (Flores et aI., 1987; Hamill et aI. , 1987; Rhodes et aI., 1987; Scheidegger, 1990; Toivonen, 1993). However, in order to reliably design and scale-up HR culture systems. their growth kinetics must be understood and mathematically mode led (Flint-Wandel et al. 1993 ; Inomata et al.. 1993; Kim et aI., 1995; Taya et aI., 1989). The shape of the HR batch growth curve has been described using Monod's equation (Hilton et aI., 1988) or the logistic equation (Toivonen et aI., 1990), both of which predict the change of total root biomass with time. However, published results (Croes et aI., 1989; Yonemitsu et aI., 1990; F1ores, 1987 ; Aird et aI., 1988) suggest that different HR morphologies result in dissimilar levels of secondary metabolite production. Thus two HR cultures with a similar biomass but different root architectures could have completely different product yields. A morphological structured model would therefore be of great interest. 8ecause IIR cultures are highl) branched and are not connected to aerial plant parts, morphological structured models of natural root systems cannot adequately describe their architecture. A kinetic model of branching growth of plant HRs published by Taya et al. (1989) assumes linear root extension, binary division of the root-tip meristem and a linear relation between biomass and root length. Although this model provides a close correlation with the biomass data of some HR cultures, its description of HR morphology and branching patterns does not conform to the findings obtained in our experimental system (Duran, 1993). HR laterals are formed in the zone behind the tip meristem, resulting in delayed generation of successive branches, as in natural plant roots (Rose. 1983). Here \\t: present a morphological structured model for HR cultures, that allows detailed description of root architecture and evaluation of root biomass at any desired moment of growth in a batch culture. EXPERIMENT AL HR cultures of Symphyflll11 officinale were maintained by subculturing a single root tip in 250 m I Erlenmeyer flasks containing 30 ml of liquid MS (Murashige and Skoog, 1962) medium with 3% sucrose. The tissue was incubated at 25°C in the dark at 150 rpm on a gyratory shaker. Every 3-4 days the fre sh and dry weights of three root systems were measured. Their morphological characteristics (length and diameter of each lateral) were determ inated by image analysis using a desktop scanner (Pan et aI., 1991) and NIH Image program. Sugar concentrations in the medium were also determined using colorimetric and enzymatic methods (Chaplin et al.) . THE MODEL Definitions I. HR systems are described h ieran.:hically in terms ll[" 10\':" .; 0[" origin of the laleraJs (\jarley, 11)70 : Hackett et aI., 1972 ; Lungley, 1973). The inoculum tip is designated as "zero-order", laterals branching off from 173