Influence of different strains of Agrobacterium rhizogenes on induction of hairy roots in Phlogacanthus Thyrsiflorus Nees
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Date
2012
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Birsa Agricultural University, Kanke, Ranchi, Jharkhand
Abstract
The present research work has been carried out with a view to standardize the protocol
for Agrobacterium rhizogenes Conn. mediated hairy root induction on Phlogacanthus
thyrsiflorus Nees, a valued medicinal plant, using two different strains of A. rhizogenes
ATCC 15834 and MTCC 532. Aseptic explants for transformation were obtained by
inoculating shoot tips of P.thyrsiflorus on MS media supplemented with BAP – 5.0 mg/l ,
AdSO4 – 50.0 mg/l and Citric Acid- 1.0 mg/l. Both the strains were evaluated for their
transformation ability and production of secondary metabolites in P.thyrsiflorus. ATCC
15834 was maintained at 28°C for 48 hrs in YEB medium supplemented with 50.0 mg/l
rifampicin. While MTCC 532 strain was grown in recommended solid medium (without
Rifampicin) at 25°C for 48 hrs. Acclimatization of bacteria culture in MS medium for 4
hrs was found effective. The explants were treated for different co-culture periods after
which those were inoculated in solid MS basal medium and kept at 25 ± 2 °C and 16 hr
light/ 8 hr dark. The percentage of hairy root induction and number of hairy roots per explant
varied with infection period.
The strain MTCC 532 performed the highest infection frequency, 93.33% with
the highest number of hairy roots, 20.00 per explant at 2 hr 45 min co-culture after 21
days. But in case of ATCC 15834 the highest transformation frequency was found to be
90.00% at 3 hr 45 min co-culture with the highest number of hairy roots, 16.1 per
explants after 21 days of bacterial infection. Among the two different strains examined in
this study, MTCC 532 was found to be the most virulent and effective both for
transformation frequency as well as number of hairy roots/ explants.
Phytochemical analysis of induced hairy roots through 2 different strains and in vivo
grown roots was done through RP-HPLC with Photo Array Detection at 225 nm. The
Phlogantholide was identified using isocratic solvent system consisting of methanol and
Milli Q water (68:32) at flow rate of 1.0 mL/min at 40o C using C18 reverse column.
Chromatogram showed distinct peak in 3 different samples at same wavelength and
almost same retention time.
Description
Influence of different strains of Agrobacterium rhizogenes on induction of hairy roots in Phlogacanthus Thyrsiflorus Nees
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