| The mineral resources in China are mainly Low-grade complex and original metal sulfide-based. The traditional mineral separation and smelting technology with high cost and environmental pollution can no longer meet the demands of the economy, however, micro-organisms through the use of mineral-metallurgical energy microorganisms, oxidative decomposition of the metal ions into the mineral solution, and further separation, extraction of metals with low cost, raw materials low demand, the process short, simple equipment, low pollution and other advantages to has become a new way to solve the problem between resource development and environmental protection.However, the bioleaching metallurgical process is slow, leaching cycle is long, lacks resistance to some heavy issues. Especially for the native complex of low-grade sulfide ore, the biological oxidation requires more energy, which requires a higher oxidation activity of the microorganisms to destroy its crystal lattice, but how to quickly identify and screen high leaching microorganisms become key factors to the problem of the slow bioleaching and the low leaching rate; on the other hand, the biological diversity of metallurgical process also determines the leaching system and function of microbial populations and the complexity of factors, effective microorganisms in the leaching system are not highly expressed. The two difficulties restrict the development of microbial metallurgy, therefore, how to strengthen the development of bacterial leaching becomes important issue in biometallurgy, It is necessary to use newer, more powerful techniques to study the bio-leaching characteristics of the microorganism itself.A. ferrooxidans is the most studied bioleaching bacteria. It could oxidize Fe2+ to Fe3+, reduced sulfur in the oxidized SO42-, are prevalent at low and middle temperatures and in the FeS2 environment (mine heap leaching system), and is a very important for industrial production and environmental protection of the chemoautotrophic bacteria, is the most commonly used bio-metallurgical strain. It is the completion of genome sequencing the only bioleaching bacteria, so that we can comprehensive study on the whole genome of A. ferrooxidans. Based on the published TIGR A. ferrooxidans ATCC 23270 sequence information,3217 oligonucleotide probes were selected. And specific human genes and plant genes were selected as negative controls and quantitative controls. With A.f bacterial genome to DNA as a positive control,58mer oligonucleotide whole genome chip were designed, which contains 3270 probe. To verify the practicality and find the best hybridization conditions, we evaluated this chip. The results show:The best temperature for the hybridization is 45?; the signal at a concentration of 100ng value has already reached saturation and test sensitivity is 5 ng, moreover, the signal value Concentration has a strong linear relationship (r2=0.992), probe found no non-target genes with non-specific interaction between the hybrid phenomenon indicating that the oligonucleotide microarray has a good specificity.We also use the whole genome micro array constructed leaching activity of different strains of gene-level analysis. Through this experimental analysis,3217 genes were found to have 967 genes specific for different strains. The remaining 2250 (70%) were shared by total A.f bacterial gene, which can be used as the criterion of whether it is A.f; Among the other 320 genes with high oxidation activity characteristics of bacteria,135 related with resistance to ferrous iron and sulfur oxidation and other functions related genes. The bacterial genes could be used as criteria to judge the metallurgical properties of A.f bacteria. Thus a " The methods of testing Acidithiobacillus ferrooxodans and its activity by microarray technology " National Standards (GB/T20929-2007) was established.For to understand how effective microorganisms in the leaching system are highly expressed, we have cloned and sequenced the obtained sequences and NCBI acidophilic micro-organisms related known sequences of the 1194 oligonucleotide probes, including 140 16S rRNA probes, functional gene probe 1054. The functional gene probe target sequences were related to carbon sequestration, nitrogen metabolism, sulfur metabolism, iron oxide, metal resistance, electron transfer and outer membrane protein-related genes. In this chip human gene were used as negative controls and quantitative controls. In addition, gene chip containing target sequences involved nearly all the present leaching systems in the natural acidic environment, the 27 known genera and 55 species of acidophilic bacteria of all gene sequences, which can effectively distinguish acidic microbial community structure of ecosystems and functional differences.This work used 16S rRNA gene-based RFLP technology to study the geographical distribution of microbial communities in AMD, and used functional genome arrays of the second generation to study its microbial community diversity in the system development and function, composition and structure; The results show that in detection of the different AMD plots, the microbial community in terms of phylogenetic diversity, functional diversity or composition and structure are different, the surrounding environmental parameters may be the main reason for this phenomenon, especially Fe, S, Ca, Mg, Zn and Cu concentrations and the pH value is the most important factor; This study provides a powerful means to understand the geographical distribution of acid mine water, diversity, composition, structure and function, meanwhile, provide a strong scientific support to the further development of bio-metallurgy technology from the waste rock and ore in the recovery of the low taste of metal ions. |
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