Isolation of plant growth promoting bacteria from torch lake sediments and their role in phytoremediation of metal contaminated soil

In this study, we isolated eight copper-resistant bacteria from Torch Lake sediment contaminated by copper mine tailings (stamp sand). Sequence analysis of gyrB and rpoD genes revealed that these organisms are closer to various Pseudomonas species. These eight bacterial isolates were also resistant to zinc, cesium, lead, arsenate and mercury. Further characterization showed that all the strains produced plant growth promoting indole-3-acetic acid (IAA), iron chelating siderophore and solubilized mineral phosphate and metals. The effect of bacterial inoculation on plant growth and copper uptake by maize (Zea mays) and sunflower (Helianthus annuus) was investigated using one of the isolates (Pseudomonas sp. TLC 6-6.5-4) with higher IAA production and phosphate and metal soubilization, which resulted in a significant increase in copper accumulation in maize and sunflower, and an increase in the total biomass of maize. Genes involved in copper resistance of Pseudomonas sp. TLC 6-6.5-4 was analyzed by transposon mutational analysis. Two copper sensitive mutants with significant reduction in copper resistance were identified: CSM1, a mutant disrupted in trp A gene (tryptophan synthase alpha subunit); CSM2, a mutant disrupted in clpA gene (ATP-dependent Clp protease). Proteomic and metabolomic analysis were performed to identify biochemical and molecular mechanisms involved in copper resistance using CSM2 due to its lower minimum inhibitory concentration compared with CSM1 and the wild type. The effect of different bacterial inoculation methods on plant growth, copper uptake and soil enzyme activities was investigated. Four different delivery methods were used including soil inoculation (before or after plant emergence), seed coating and root dipping. Soil inoculation before sowing seeds and coating seeds with PGPB led to better growth of maize, higher copper uptake and an increase in soil invertase and dehydrogenase activities. Proteomic and metabolomic analyses were performed to investigate the effect of bacterial inoculation on maize grown in normal soil and stamp sand. Our results revealed that bacterial inoculation led to environment-dependent effects on maize proteome and metabolome.

Cloning and characterization of the genes involved in cambial growth in response to elevated [CO2]

Aspen (Populus tremuloides) trees growing under elevated [CO2] at a free-air CO2 enrichment (FACE) site have produced significantly more biomass compared to control trees. The molecular mechanisms underlying the observed increase in biomass productivity was investigated by producing transcriptomic profiles of the vascular cambium zone (VCZ) and leaves, followed by a comparative study to identify genes and pathways that showed significant changes following long-term exposure to elevated [CO2]. This study is mainly to verify if genetic modification of a few selected candidate genes including CAP1, CKX6, and ASML2 that are expressed in vascular cambium in response to elevated [CO2] can cause the changes in plant growth and development. To this end, these three genes were cloned into both sense and antisense constructs. Then antisense and sense transgenic lines of above-mentioned genes were developed. 15 events were generated for 5 constructs, which were confirmed with regular PCR and RT-PCR. Confirmed plants were planted in greenhouse for growth and phenotypic characterization. The expression of CAP1, CKX6 and ASML2 in antisense plants was measured by real-time RT-PCR, and the changes caused by gene interference in cambial growth were studies by analyzing the microscopic sections made from the antisense transgenic plants. It has been found that 1) CAP1 is mainly expressed in xylem and root. 2) RNAi suppression of CAP1 significantly affected height and diameter. 3) CAP1, ASML2 and CKX6 affected xylem and phloem cell proliferation and elongation. Due to the delay in regenerating sense transgenic plants, the characterization of sense transgenic plants is limited to growth only.