Page 116 - Plant-Based Remediation Processes

6 Metal Remediation via In Vitro Root Cultures 105

direct the production of opines. These compounds are synthesized and excreted by
the transformed cells and consumed by Agrobacterium as a nutrient source
(Dessaux et al. 1992). According to the opines produced, the Ri plasmids can be
classified into several lines: octopine, agropine, nopaline, mannopine, and the
stereoisomers cucumopine/mikimopine (Hu and Du 2006; Veena and Taylor
2007). The T-DNA is defined by two border sequences from 25 bp in length and
highly homologous in sequence. The T-DNA contains several genes, some of them
involved in auxin biosynthesis and sensitivity (aux), which cause differences in
hairy root growth and morphology when compared to non-transformed roots
(Meyer et al. 2000; Christey 2001; Chandra 2011). The genes aux1 and aux2
seem to be responsible for auxin autotrophy of transformed roots. The genes rol,
particularly rol A-B-C-D genes, affect the development of the infected plant cells,
inducing the “hairy root” syndrome, the rolB gene being the most important. The
product of rolC influences the metabolism of cytokinins and gibberellins (Estruch
et al. 1991; Nilsson et al. 1993) while the rolB protein enhances auxin binding to the
cell membrane (Filippini et al. 1994; Veena and Taylor 2007). For a more detailed
description of the rhizogenic process, see Chandra (2011).
The extensive root proliferation induced by A. rhizogenes, generally considered
an undesirable characteristic, may find good utility for phytoremediation due to the
larger root surface to uptake the contaminant from the medium. Transformed root
cultures present faster growth than non-transformed roots, are genetic and biochem-
ical stable, and have hormonal autotrophy, that is, roots have the unique ability to
grow in vitro without PGR and then are easily established and propagated in the
laboratory (Shanks and Morgan 1999; Suza et al. 2008).
Root hair formation occurs within a specific region of the root, a short distance
above the region of root elongation. Root hairs are short and develop on both
primary and secondary roots. Interestingly, a root hair is a single cell that consists of
a thin cell wall, a thin lining of cytoplasm that contains the nucleus, and a large
vacuole containing cell sap (Gillaspy 2008). The hairy root disease is characterized
by plagiotropic root growth, a high degree of lateral branching, and the profusion of
root hairs, although the tissue maintains a highly differentiated and functional root
organ. These roots also present a higher enzymatic degradation capacity due to the
peroxidase, laccase, and oxygenase content (Flocco et al. 1998; Boominathan and
Doran 2003b; Talano et al. 2003; Telke et al. 2011). The transformed roots offer the
interesting property that the whole plant can be easily regenerated (Tepfer 1990;
Giri and Narasu 2000). The hairy root cultures have proved to be successful in vitro
systems for studying the phytoremediation process. Table 6.1 shows some
examples of hairy root cultures used for metal removal.

6.5 Mechanisms of Tolerance to Heavy Metals Present on Hairy
Root Cultures

The employment of hairy roots has been focussed on the extraction of Cd, Ni, U,
Zn, and Cu. The mechanisms of tolerance of these metals are discussed below.