Characteristics of Heavy Metal uptake and Accumulation in Selected plant Species for Development of Phytoextraction Technology.

Abstract

soil, especially agricultural fields contaminated with toxic metals, has become a global problem and demands economic, efficient and environment friendly remediation technologies. Phytoextraction is a potential plant-based technology for the decontamination of polluted soil and water. It is an economic, solar driven, and environment friendly technology. In the present study, physiological, biochemical and molecular characteristics of cadmium uptake and accumulation in three plant species (Parthenium hysterophorus, Ricinus communis and Cannabis sativa) were studied for the development of phytoextraction technology. In the first experiment, different treatments of Gibberellic Acid (GA3 10−2, 10−4 and 10−6 M as foliar spray) and Ethylenediaminetetraacetic acid (EDTA 40 mgKg-1 soil as single dose and 10 mgKg-1 soil given in four doses) were studied for their effects on Cd phytoextraction, and concentration of proline, phenolics and chlorophyll in Parthenium hysterophorus plants grown in Cd (100 ppm) contaminated soil. The plants showed Cd hyperaccumulator potential based on Cd bio-concentration factor (BCF > 1 in control plants). The GA3 and EDTA application increased the extent of Cd phytoextraction by the plants. Most significantly increase in Cd accumulation and bio concentration (BCF 9.75 ± 0.34) were found in plants treated with GA3 (10−2 M) in combination with split doses of EDTA. Gibberellic acid significantly increased the concentrations of phenolics and chlorophyll in the plants. Cadmium accumulation in plant tissues showed positive correlation with free proline (R2 = 0.527, R2= 0.630) and total phenolics (R2 = 0.554, R2 = 0.723) in roots and leaves, respectively. In the second experiment, physiological and biochemical analyses were performed. The objectives were to assess the effect of Molybdenum (Mo 0.5, 1.00 and 2.00 ppm) on Cd phytoextraction, and concentration of endogenous proline, phenolics and photosynthetic in Ricinus communis and Cannabis sativa plants grown in Cd (25, 50 and 100 ppm) contaminated soil. Molybdenum was applied as a foliar spray, soil addition and seed soaking. Foliar spray of Molybdenum highly increased Cd uptake and accumulation in both plants. Molybdenum seed soaking and foliar spray highly increased the biomass, concentration of free proline and total phenolics as compared to control plants. Positive correlations of proline and phenolics with Cd accumulation were found in roots and leaves; suggesting a significant role of proline and phenolics in Cd phytoaccumulation. Molecular investigation was carried out with objectives: (1) To determine the presence and then expression of DREB-1A, DREB-1B, DREB-1F and CBF like genes in Ricinus communis and Cannabis sativa plants, (2) To evaluate the effect of molybdenum and cadmium on expression of these genes, (3) To correlate the expression of genes with Cd accumulation, and free proline and total phenolics concentrations in plants. Molybdenum was applied as a foliar spray (0.5, 1, 2 ppm) while Cd (50 ppm) was added to soil. cDNA was synthesized through reverse transcriptase Polymerase chain reaction (RT-PCR). PCR from genomic DNA and cDNA with genes specific primers were performed. The PCR products were sequenced and compared the nucleotide sequences and deduced amino acid sequences for homology with other plants. Results confirmed the presence of DREB-1A, DREB-1B, DREB-1F and CBF like genes in R. communis. In C. sativa the genes were identified for the first time. Cadmium induced the expression of DREB-1B, DREB-1F and CBF like genes in both the plants while molybdenum foliar spray further increased the expression of these genes under Cd stress. The DREB-1A showed no expression in both the plants while its presence was confirmed by the PCR product of genomic DNA with gene specific primers. The CBF-like gene was expressed in both the plants sprayed with molybdenum (without Cd), while the other genes were not expressed with Mo only. The DREB-1F and CBF like gene of both the plants showed more than 80 % nucleotide sequence homology with these genes in other plants. Predicted amino acid sequence of DREB-1F from both plant showed more than 75% homology with protein sequences of other plants, while the CBF like gene demonstrated more than 80% homology with protein sequences of other plants species. The expression of DREB-1B, DREB-1F and CBF-like genes (semi quantitative) was positively correlated with Cd accumulation, free proline and total phenolics in Cannabis sativa plant, while these correlations were significantly positive in Ricinus communis. Application of GA3 and Mo increased concentration of Cd in plants tissues. A positive inter-correlation was found between Cd concentration in plant tissues, production of prolin and phenolics and the expression of DREB-1B, DREB-1F and CBF-like genes.