21-12-2012, 06:27 PM
HAIRY ROOT CULTURES AND THEIR ROLE IN SECONDARY METABOLITE PRODUCTION
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INTRODUCTION
Hairy root culture , also called transformed root culture , is a type of plant tissue culture that is used to study plant metabolic processes or to produce valuable secondary metabolites ,often with plant genetic engineering.
Hairy root cultures can be useful especially for the production of root-associated metabolites because of their high-growth rate and genetic stability.
Establishment of hairy root cultures by genetic transformation of plant tissue by the pathgenic bacterium Agrobactium rhizogenes.
GENETICALLY TRANSFORMED CULTURES
The Ri plasmids can be engineered to also contain T-DNA used for genetic transformation of the plant cell.
The resulting genetically transformed root cultures can produced high levels of secondary metabolites , comparable or even higher than those of intact plants.
Three genetic elements are known to be essential for the control and regulation of successful transfer of the T-DNA from Ti or Ri plasmids to plant cells.
The right T-DNA border sequence
The plasmid vir genes and
Three bacterial chromosomal vir genes
MARKERS TO CONFIRM TRANSFORMATION
Confirmation that a plant cell is transformed on infection with Agrobacterium rhizogenes can be obtained in several different ways using markers:
Morphological marker :
Roots transformed with A.rhizogenes have an altered phenotype and these roots with their profusion of laterals and lack of geotropism grow readily in culture.
Also , the transformed regenerants of hairy roots inherit an aberrant phenotype in having wrinkled leaves , shortened inter node and several other morphologically distinct features compared to their normal counterparts.
Growth characteristics of the established root clones :
Doubling time :
In transformed root cultures , an exponential increase in biomass occurs as a result of lateral root formation and a consequent exponential increase in the number of elongating tips.
In 1988 , Flores et al. have reported an average doubling time for different hairy root cultures of family Asteraceae and shown it to be rarely more than 24 hours , explaining higher growth rate associated with hairy roots tissues in culture.
An average doubling time for cultured hairy roots of Hyoscyamus species is reported to be between 40-60 hours.