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    • 专利摘要:
    This invention corresponds to a bacterial cell or preferably a yeast cell, which in its natural state exhibits a reduced tolerance to iron and sulfur but a high capacity to accumulate phosphorus in the form of polyphosphates. When this cell is subjected to a process of mutation by its own endogenous mechanism of iron transport (protein complex [FTR1 / FET3]), it acquires a high tolerance to iron. In addition, the cell of the invention tolerates and accumulates sulfur when it is genetically transformed with a vector that allows expression of the sull gene encoding a high affinity sulfate permease (SUL1). The cell of the invention also increases its natural ability to accumulate phosphorus in the form of polyphosphate when genetically transformed with a vector that allows expression of the pho84 gene encoding a high affinity phosphate transporter (PH084). The mutated and double-transformed cell described according to the invention has a high capacity to remove phosphorus and / or sulfur from liquid or solid culture media, even if these media correspond to iron ores or substrates with more than 50% by weight of iron.
    公开号:SE1300669A1
    申请号:SE1300669
    申请日:2013-10-21
    公开日:2013-11-14
    发明作者:Viviana Rosa Ordenes Ortiz；Darwin J Mutarello Wuillans；Nia C Oetiker Mancilla
    申请人:Brotar Limitada；
    IPC主号:
    专利说明:

    Since its content of impurities is high, a process called pelletization is carried out. This is the case for the Los Colorados and Algarrobo mines, where the mineral is transported to the pellet factory in the Huasco Valley.
    Here, the material is reduced in size in a process called grinding and grading; it is machined by rotor mills which reduce the diameter of the material to 44 μm and separate it by magnetic means (2).
    Phosphorus in concentrations between 0.08% and 0.12% is easily processed in steelmaking, but concentrations above this level are detrimental to production because phosphorus is an element that increases the fluidity of molten metal and reduces the melting temperature. As for sulfur, it is considered a harmful substance in iron alloys, a pollutant. But sometimes 0.25% sulfur is added to improve the machinability of the iron.
    High sulfur steels are difficult to weld because they cause porosity in the weld melt, and this is the reason why one wants to remove this contaminant completely during the extraction process (3). There are deposits with enormously high levels of pollutants in the end product. In the second region's (Antofagasta) largest mountain range we find the El Laco deposit. According to Compañia Minera Del Pacífico, it has a reserve of more than 224 million tonnes of minerals. It consists mainly of magnetite. At present, this deposit has not yet been exploited because the extraction processes have shown that this mineral contains 2% phosphorus (whose tolerance limit is 0.1%), which means that the product is not commercially viable (4).
    Phosphorus in the form of orthophosphates (Pi) is considered an impurity in the process of producing iron, but it is of great importance for all living organisms and accumulates inside the cells in the form of polyphosphates (polyP). Polyphosphates are linear polymers that contain hundreds of orthophosphate residues connected via high-energy phosphate bonds. This can be found in cells in free form or in complex states with ions such as calcium and magnesium, certain amino acids (lysine, arginine), proteins and nucleic acids. In microbial cells, it is a multifunctional component: 1) It is a reserve of large amounts of intracellular Pi and regulates their levels. 2) It acts as an energy reserve as the polyP-phosphanohydride bonds have energy in hydrolysis similar to ATP, and in addition synthesis of ATP from polyP in vacuoles has been demonstrated. 3) In lower eukaryotes, capture and storage of cations in vacuoles is observed, where polyP can enclose them in an inert osmotic manner. 4) On the other hand, a polyP-Cah-polyhydroxybutyrate complex can be formed with calcium, and it acts as a transmembrane channel and participates in the cation transport process. 5) PolyP is found in the cell walls of lower eukaryotes, where they are very important in maintaining the negative charge of the fungal surface, which has been shown to enable interaction between cells and between fungus and plant. 6) As a polyanion, it can interact with proteins and enzymes with particularly high levels of acid residues, and can therefore modulate its activity whether it is in competition with the substrate for binding or through its interaction with polycationic activators. 7) Finally, it regulates the enzyme activity and expression of a large group of genes that form the basis of its survival mechanism under stressful conditions. (5) For example, in Duna / iella salina, where the alkalization of the cytoplasm is the result of a significant hydrolysis of polyP due to a major exopolyphosphate synthesis which hydrolyzes the terminal residues of polyP to release Pi, resulting in in pH stabilization. This is why it is considered to play an important role in increasing resilience under adverse environmental conditions. Another example of this is the accumulation of polyP in Escherichia coli under osmotic and nutritional stress, or Acinetobacter spp which accumulates polyP when subjected to alternating aerobic-anaerobic cycles. (6) The increase in intracellular polyP levels is due to the stimulation of its uptake from the extracellular environment, which is performed by the high affinity PHO84, H * / PO 4 (3 ') symporter. Cryptococcus humico / us, an Antarctic yeast fungus that grows above ground in soil of volcanic origin, has the capacity to accumulate 22 mg / l polyP when exposed to pH 5.5, which is 10 times more than the accumulation of polyP at normal pH levels (7.5). (7) This yeast has been used in phosphate elimination processes from wastewater to regulate the eutrophication process. Doctors John Quinn and John McGrath, both from Northern Ireland, have, together with Northern Ireland Water Service, Severn Trent Water Ltd, Extract Solutions Ltd and Yorkshire Water Ltd, evaluated the use of this yeast in water purification in a large-scale pilot plant. The results show that they achieved an 80% elimination of phosphate from the eutrophically treated water. Unlike what was done with wastewater biosanification, if C.humico / a cells are used to remove phosphate from iron concentrate (eg from materials in the El Laco deposit), these cells will be exposed to high iron concentrations. , which could generate different cellular responses, which can range from a high iron uptake to intolerance and on to subsequent cell death (8). On the other hand, sulfur is an essential macronutrient for animals and plants. It has been neglected to study it for many years (compared to other essential nutrients).
    However, it is very important in our environment. At least 95% of the sulfur in the soil is in organic form. Exceptions to this occur in limited areas where salty soils, acid sulphate and other things have a high content of sulfur-containing minerals. Transformations of the sulfur in the soil are mainly determined by the activity of microorganisms and can be classified as mineralization (from organic sulfur to sulphate mineral), immobilization (conversion of mineral sulphate to organic sulfur), oxidation and reduction (conversion of inorganic sulfur to low or high oxidation state compounds) ( 9).
    Sulfate is one of the most common anion macronutrients after phosphate in cells and is the largest source of sulfur for many organisms. Like all nutrients, sulphate is taken up and accumulated in the cell by a very specific transport mechanism on the plasma membrane (10). Some organisms such as metazos or parasitic microorganisms depend on the organic sulfur found in amino acids, while plants, fungi and bacteria assimilate inorganic sulfur. Most of them take in sulfur in the form of sulphate, reduce it and incorporate it into cysteine and homocysteine. Assimilation of sulphate takes place in five enzymatic steps. First, it is activated to adenylyl sulfate (APS) and 3'-phosphoadenylyl sulfate (PAPS) by ATP sulfurylase and APS kinase. PAPS is then reduced by PAPS reductase to sulphite, and sulphite is reduced by sulpheductase to sulphide. Finally, cysteine is formed when sulfide of OAS- (thiol) lyase is incorporated into O-acetyl-L-serine (OAS). The enzyme 3'-phosphoadenylyl sulfate (PAPS) reductase catalyzes the first reducing step in this sequence. It uses thioredoxin (Trx) or glutaredoxin (Grx) as an in vitro hydrogen donor to reduce 3-phosphoadenylyl sulfate to free sulfite.
    Sulfate uptake is a tightly regulated process that controls the homeostasis of sulfur inside the cell. This is regulated by the organism's nutritional status and the external availability of the nutrient. Lack of sulfur can result in high uptake rates of plants, bacteria, yeasts and filamentous fungi.
    However, sulfate uptake decreases when sulfur-containing compounds, such as glutathione, cysteine or H25, are available (11). In plants and certain microorganisms, sulfur is required for the synthesis of various compounds, including cysteine and methionine, proteins, glutathione and secondary metabolites such as glucosinolates. This requirement is the reason why sulfur is considered a macronutrient for growth, development and reaction to both biotic and abiotic stress. The uptake and transport of sulphate is energy dependent (and driven by a proton gradient generated by ATPases) through a proton / sulphate transport. In plants, after absorption through the plasma membrane of cortical root cells and epidermal cells, the sulphate is transported into the plant through cell membranes under the control of a set of sulphate transporters. Sequencing of the Arabidopsis tha / iana genome led to the identification of a multigen family (14 genes) encoding what are thought to be sulfate transporters (12). In Neurospora / Aspergia / Ius, yeasts and other fungi as well as in plants, inorganic sulphate is used in a similar way. After cell uptake, inorganic sulfate is phosphorylated with adenosine triphosphate (ATP) in two enzymatic steps to generate PAPS, reduced to sulfite and then to sulfide, which condenses to o-acetylserine to form cysteine, which acts as an intermediate in the synthesis of methionine and S- adenosylmethionine, as mentioned above (13). In eukaryotes, SulP proteins have largely the main responsibility for sulfate uptake. These proteins are anionic transporters or anion anion exchangers. Many of them have been functionally characterized and differ in their affinity for substrates. Some of them act as Hïsulphate imports or sulphate / bicarbonate: Naïsymports. These proteins have a common topology of 12 transmembrane helices in the sulfate transport domain. Domains have positively charged arginine in extracellular loops that play an important role in sulfate capture (14).
    Studies of sulphate transport in fungi have been limited to a small number of species, e.g.
    Saccharomyces cerevisiae, Neurospora crassa and Penici / Iium chrysogenum. Two sulphate permeases belonging to the Su | P family have been identified in these species. In yeast, transcripts of two permease genes, SUL2 and SUL1, have been characterized. Both permeases have 12 transmembrane domains and appear to be very similar. Kinetic studies have reported that both have a high affinity for sulfate transport (14). On the other hand, iron (Fe) is an element required by almost all organisms; its biological significance depends on its chemical form, which varies between divalent (ferrous) and trivalent (ferric) due to oxidation / reduction (redox) reactions. This chemistry is used in vital physiological activities such as oxygen transport using hemoglobin and production of ATP by aerobic cell landing. Unfortunately, the same redox reaction helps to convert superoxide and hydrogen peroxide into powerful oxidizing hydroxyl radicals that trigger cell damage (Fenton reaction) (15). Previous work has experimentally demonstrated iron toxicity in yeasts: cells were exposed to high concentrations of iron salts [20 mM] for 4 hours resulting in a low survival rate under aerobic conditions (16). In yeast, iron homeostasis is the balance between taking iron for growth and cell division in the context of meeting the demands of all cells. Fungi maintain this balance through post-transcriptional regulation of genes encoding proteins that use iron that are involved in storage and recycling. All fungi examined in detail use at least two of the four major transmembrane mechanisms for iron uptake. One of these mechanisms is mediated by siderophores (17). Although only some yeasts can produce their own siderophores, all of them can express siderophor receptors. Siderophores have the main purpose of solubilizing the iron which is generally insoluble in the ferric form by the action of a reductase. S. cerevisiae produces at least seven reductases encoded by genes FRE1-7. In contrast to siderophores, this mechanism involves the direct entry of ferrous ions, and it is used only for a few yeasts, such as Saccharomyces cerevisiae, which have Fet4 as a carrier. Orthologs of Fet4 have also been found in the genomes of Schizosaccharomyces pombe (NP595134.1), Candida glabrata (XP_445982.1) and Kluyveromyces lactis. A third mechanism is by means of high affinity iron transport systems comprising a multicopper oxidase, which catalyzes the reaction to oxidize four moles of Fe Characterized by yeast and appears to be a conserved system It has been determined that the Km for Error * of this transport system is about 0.2 μM, a value indicating an efficient system for the accumulation of iron.This system has a higher affinity than other iron transport mechanisms, with 10-100 times better kinetics.It has been determined that the iron concentration in a cell is about 60 μM, at a cell volume of 60 μM.This concentration is largely maintained by the action of the high affinity complex for Fe transport [Fet3 / Ftr1] because it has documented that degradation of the gene encoding Ftrl protein results in a 95% reduction in iron uptake without affecting the growth of the yeast (18).
    Detailed description of the invention Based on what is described above, we have concluded that yeasts of the species Cryptococcus humicolus can be used to remove phosphate and sulfur from iron ore (due to their high capacity to accumulate polyP). Nevertheless, the large accumulation of Fe (which is taken up by the yeast cells that grow iron ores) can cause cell damage with concomitant ion imbalance leading to cell death.
    Therefore, a first step was to provide C. humicolus with high iron tolerance (250% by weight) using genetic engineering. For this, the yeast cell was subjected to a mutation process by the endogenous mechanism of iron transport (complex [FTR1 / FET3]). Physiological and mineralogical studies of C. humicolus allowed us to confirm that the mutation of [FTR1 / FET3] caused increased iron tolerance. For phosphorus removal, the natural ability of this yeast fungus to accumulate phosphorus as polyphosphate (polyP) (7) increased with genetic transformation that enabled the introduction and expression of the gene pho84 from Saccharomyces cerevisiae, which encodes the high affinity phosphate transporter PHO84. In addition, to remove sulfur, the yeast fungus Cryptococcus humico / us, which had already been transformed with pho84, was again transformed with the gene sull from Saccharomyces cerevisiae, which encodes the sulphate permease SUL1. This second transformation gives the described cell the ability to accumulate sulfur contents. The described cell has the ability to express both the foreign genes, which give the transformed cell a much better ability to capture and accumulate sulfur and phosphorus from aqueous or solid media with high concentrations of these minerals. In addition, mutation of high-affinity iron transport systems allows the cell to grow and multiply rapidly in high iron media (250% by weight). Our invention relates to a bacterium or a yeast which is preferably 1) transformed with the Saccharomyces cerevisiae gene sull, which encodes the sulphate permease SUL1, which gives the transformed cell increased tolerance to grow in media with high concentrations of sulfur, and the ability to accumulate the substance inside the cell 2) is transformed with the Saccharomyces cerevisiae gene pho84, which encodes the phosphate transporter PHO84 with high affinity, which increases the natural ability of C. humicola cells to accumulate phosphorus as a polyP and 3) is subjected to a mutation process by the endogenous mechanism of iron transport (complex [FTR1 / FET3]), which gives it the ability to grow and multiply in iron-concentrated medium. The double-transformed cell, simultaneously mutated according to the invention to be grown in an aqueous or solid medium with high levels of sulfur, phosphorus and iron, can thus tolerate high iron concentrations and at the same time remove a significant part of the sulfur and / or phosphorus. These functions have been conferred by the genetic modifications set forth above. This cell could be used as a technical innovation for iron ore mining as it enables direct removal of impurities such as phosphorus and sulfur from iron ore. This invention therefore provides an alternative for removing phosphorus and sulfur from iron ore.
    The purification process for the ore has hitherto been carried out in an inefficient manner by physical removal consisting in reduction of the particle size or pelletization. Pelleting does not work in all individual cases to remove contaminants from iron ore. An example is the El Laco iron deposit in northern Chile, where mechanical removal is not effective in removing phosphorus and sulfur, so consequently the mineral cannot be used. The scope of the present invention offers a possible use for removing contaminants from iron ore at low operating cost. In addition, the described invention also finds application in decontamination of wastewater and / or contaminated industrial sludge, with high levels of substances such as sulfur, phosphorus and / or iron, where the cell according to the invention can contribute to decontamination and / or recovery of the minerals mentioned above. Exemplary of the Invention We describe below examples to reproduce the present invention and the results we have obtained, which show the great advantage of genetically modified yeast according to the invention for bioaccumulating sulfur and phosphorus from solid or liquid media, especially from them. which have a high iron content (over 50% by weight). These examples are intended only to illustrate the invention without limiting it, as one skilled in the art will appreciate that it is possible to broaden their scope. What is shown here is the potential for the genetically engineered cell of the invention, which can promote either the removal of contaminants from iron ore or the purification of contaminated water from industrial processes.
    Example Example 1.
    Increasing the capacity of C. humicolus to accumulate polyphosphate: Analysis of natural polyP accumulation in C. humicolus, which is achieved by culturing yeast at pH 5.5, and also at pH 7.5 in different potassium sulphate concentrations. The analysis of the po | yP accumulation is achieved by means of a colorimetric method using ammonium molybdate in acidic medium; phosphate accumulation is verified under a microscope. Obtaining a yeast capable of hyperaccumulating polyphosphate (polyP) by overexpression of PHO84, a sulfate permease from Saccharomyces cerevisiae. The transformation of Candida humícola was performed with the sequence codings for sulfate permease (PHO84). This transporter is found in the plasma membrane of fungi, and its function is the transport of inorganic phosphate (Pi). This sequence was incorporated by means of a genome integration system, which includes several copies of the gene into the yeast genome, with overexpression of this transporter and promotion of Pi uptake as a result. Analysis of polyP accumulation in genetically modified C. humicolus, which is achieved by culturing yeast at pH 5.5, and also at pH 7.5 in different potassium sulphate concentrations. The analysis of the polyP accumulation is achieved by means of a colorimetric method using ammonium molybdate in acidic media, and the accumulation is verified under a microscope.
    Example 2.
    Obtaining a yeast that can accumulate sulphate at the intracellular level by its removal from the environment in which it grows: Analysis of natural sulphate accumulation in C. humicolus, which is achieved by culturing yeast at pH 5.5, and also at pH 7, 5 in different sulphate concentrations. The analysis of sulfate accumulation is achieved by a methylene blue method after basic oxidation using NaOBr, which is widely used in the analysis of intracellular sulfates in plants, as well as the use of cell incubation in [3sS1 sulfate and post-uptake analysis by radioactivity measurements. Obtaining a yeast that can accumulate sulfate by overexpression of the cell membrane permease SUL1. The transformation of Candida humicola was performed with the sequence encoding a high affinity sulfate permease on the plasma membrane. This sequence was incorporated by means of a genome integration system, which includes several copies of the gene into the yeast genome, with overexpression of this enzyme and increase of the yeast sulphate uptake as a result. Analysis of sulphate accumulation in genetically modified C. humico / us, which is achieved by culturing yeast at pH 5.5, and also at pH 7.5 in different sulphate concentrations.
    The analysis of sulphate accumulation is achieved by a methylene blue method after basic oxidation using NaOBr, which is widely used in the analysis of intracellular sulphates in plants, as well as the use of cell incubation in [35S] sulphate and post-uptake analysis by radioactivity measurements.
    Example 3.
    Obtaining C. humico / us yeasts that can survive at high concentrations of iron (Fe) and with a minimal capacity for metal accumulation: Analysis of natural tolerance of C. humicolus at high concentrations of iron, by growing yeasts at high concentrations iron concentrations at pH 5.5 and at pH control (7.5). For the analysis of intracellular (ICP-AES).
    Obtaining yeast that can survive at high Fe concentrations. To achieve this, iron accumulation was performed using inductively coupled plasma atom absorption spectroscopy using a mutation of the ftrl gene because it encodes the iron permease of the yeast fungal plasma membrane in C. humicola. First, an analysis of the gene with bioinformatics (CLUSTAL.X, BLAST) was performed in order to be able to carry out an adjustment of the sequence encoding Ftr1 for different yeasts that are phylogenetically related to C. humicola. Subsequently, degenerate primers were generated for gene amplification. The amplified fragments were sequenced (TAKAHASHI et al). Based on accurate knowledge of the Ftr1 sequence in C. humicola, precise primers for its amplification were constructed by RACE PCR, thus obtaining the terminal 5 'and 3' fragments of the sequence. By determining these terminals, a 50 bp amplification was made for each 5 'and 3' gene terminal with restriction sites to bind these fragments to the pUG6 vector. This vector is based on the Cre-loxP gene disassembly system. It has its own P1 bacteriophage fragment for loxP whose genome contains recombination sites consisting of 13 bp (repeated-inverted, loxP) with a gap of 8 bp. These places are recognized by an enzyme called Cre. Each of the loxP sites is bound by two recombinase molecules. The Cre-IoxP complex is combined with another complex present in the DNA molecule, and an asymmetric cross-section is made of the recombinase which results in a division of the DNA region between the two sites. The pUGG vector contains the loxP-kanMX-loxP module, and the gene fragment to be mutated is added on both sides of the loxP sites. When the vector is introduced into the yeast fungus, a breakdown of the Ftr1 gene takes place, which is checked by PCR technique. Assessment of the ability of C. humicolus Aftrl to bioaccumulate polyP and grow at high Fe concentrations. At this stage, the growth of the mutated yeast was assessed at pH 5.5 and 7.5 was evaluated, and this growth at high iron concentrations and its accumulation was analyzed by ICP-AES.
    Description of fi gures Fig. No. 1 Schematic illustration of polyP accumulation in Cryptococcus humicolus (or Candida humicola). Pho-84 is a high-affinity transporter on the plasma membrane, which together with pyrophosphokinase enzyme (PPK) regulates the cell's level of phosphate (N = core, PPK = pyrophosphokinase, V = vacuole, polyP = polyphosphate).
    Fig. No. 2 Schematic illustration of the strategy used to increase the capacity of Cryptococcus humicolus (or Candida humicola) to accumulate polyP. This was done by transforming yeast cells with Pho-84 using a system for stable transformation of yeast (P = promoter, N = nucleus, PPK = pyrophosphokinase, polyP = polyphosphates, V = vacuole).
    Fig. No. 3 Hypothetical model representing sulfate uptake in Cryptococcus humicolus (or Candida humicola). X is the putative sulfate permease of C. humicola (N = core, S = sulfate, X = sulfate permease).
    Fig. No. 4 Schematic illustration of the strategy used to increase the capacity of Cryptococcus humicolus (or Candida humicola) to accumulate sulfur. This was done by transformation with SUL1 using a system for stable transformation of yeasts (P = promoter, N = core, SUL1 = sulfate permeate of Saccharomyces cerevisiae, X = SUL1 ortholog in C. humicola, S = sulfate).
    Fig. No. 5 Schematic illustration of iron uptake of Candida humicola by high affinity iron transport complex Fet3 / Ftr1 (N = core, FTR1 = high affinity iron permease, FET3 = ferro-Oz oxidoreductase, Fe = iron). Fig. No. 6 Schematic illustration of the strategy used to increase the iron tolerance of Cryptococcus humíco / us (or Candida humicola). This was done by deletion of the gene encoding Ftrl, a part of the high affinity iron transport complex. The mutation was performed using the Cre-loxP system (N = nucleus, Cre-loxP = system for deletion or decomposition of genes, FTR1 = high affinity iron permease, FET3 = ferro-Oz oxidoreductase, Fe = iron).
    Fig. No. 7 Phosphorus removing activity measured in Cryptococcus humicola cells transformed with PHO84 and cultured under 72 hours of culture medium with high iron concentration (50% by weight). The cells were compared with untransformed yeast grown under the same conditions.
    Fig. No. 8 Sulfur removal activity measured in Cryptococcus humico / α cells transformed with SUL1 and cultured for 72 hours in culture medium with high iron concentration (50% by weight). The cells were compared with untransformed yeast grown under the same conditions.
    Table No. 1 Generation times (Gt) for C. humicolus strains mutated the high affinity iron transport system FTR1 / FET3. The generation time (Gt) was measured during the exponential growth phase in culture medium supplemented with iron, 50% by weight. The exponential growth rate of yeast cultures is expressed as generation time in hours (h), i.e. the doubling time for each yeast fungus population tested.
    The yeast mutants were compared with the control population without mutation.
    Nucleotide Sequence No. 1: DNA encoding the high affinity FTR1 (iron permeao) component of high protein protein complexes [FTR1 / FET3].
    GTCTFCTGTTCTGCCTITFAACTCTFTGGACAAAAGTCCAAGTACTTCAAGTCGACCGGTC GACTCTGÛTFTTATTCTITCCCACTFG CATCGAAGAGCTGTTCTGTAACATCCCTGACCC AAGACAGACTCT G CCACCATGGCGAAAGACGTFITCTCAGTACCCATCTTCTTCATCATCT TCCGTAAGTAGTATITITITTGGTCCTTCATTACTCTÜTACACCCACATCAGATITACTCA CG CTAATCTCCCACAATAGGTGAGACGGTTGAAGCTG CTATCATTGTCTCCGTCCTTCTTT 10 10 15 20 25 CTITCGTCGAACAG CTTATG CTCCATGGTTCACTCG CCCAGGACGAGAACGACCTCAGTA ATGAAAACGCCTCCAACAATGGATCAG CTGGGGACG CAGAAGCCCCAACGAACTCTATT ACCGACAAGGAG CGACGAG CAAAGTTAGTTAAGAGGATGAGGATTCAAATCTGGGCGG GTACAGGTGTCGGTTTCFTCATTG CTATITGTATTGGTG CTG CTTTTATCG CTGTCGTATG TTTCCCATHTFGTAACTACATCCGAGGAGCGACG CTAAACTATGATITACG CCAGTITFA CACCACACTCAACGATCTATGGGCCGACACTGAACAGATITGGGAAGGTGTCTTCTCCGT CATCG CTG CTGGTATCATCTACGTATGTTCTGTCCCCACTTCCCAACTTTTATTGATACTAA CACACTTCCTITCTCCTAG CTAATGGCCACTGCCTFCCTCAAAATGGATAGAAG CCGTATT AAGTGGCGATGGAAG CTTG CCGCCG CTTTCGACCGAAGCCAAG CCAAATTG CTCGCCCG TGAAAAGATGTCTGAACACGACAG Gaag CTTG CAGAGAAGGAAGGCAAGAG CG GAAA ATGGGCGTTATTCCTTCTTCCGTrCATCACTGTGTTG CGTGAAGGTCTTGAGGCTGTFGTC 1TTGTCGGTGGTGTGAGTGACTITITCTCTFTCTCAATATCATTTAGATGGACAGGATTGA TGATGAGAAAG GTGTCCCTFG GTATTCCAG CT ACTTCTATCCCG CTCG CCGTCGTTGTTG GCAWATFGCCG GWTCGCCGTFGGTAAGTCGTCG CTTITCCTCTFFCACTFCCCAG CG CA TAGTG CTCACCATCAACCCGGCACTCAAAAGGCTACTTGATCTATCG CACCGGCTCTACC ACCACCCTTCATTGGTTCCTCATCGGTTCCACCTCTITCCTCTGCCTTATCGGGGCCGGTC TTCTGTCCAAAGGTGTCGGCTTCTTCCAATACTACCGTITCGCCAAAGGTGTGGGTGGGG ATGTAG CAGAGACTGGAGATG GGCCTGGTTCGTTCCÅG GTCG CAG GG AATGTTTG GCA TTTGGAGTATGGAAACCCAGAGGTCAGTCGTTATCTCATAAGATCAAAGATCTTTCACTA CTTTCAAAAGGGTATGTGTG CTAATGTCUTCATGTAGACTG GTTCTGCTACGACTAACG GTGGTTGGCAAATCTITAATG CCATITTCGGCTGGAGTAAGTFCTCCCTTCTCCTAACCCC GTACATG CAGATTACTAACCACACCCCACCTCTTAATITCTCATAGACAATACGGCCACCC TG GG CÅCTATCCTCTCTTACGTGTTCTACTG GATCCTCGTCATFG CCACCCTG ATCTATCT CAAATGGAAGGAAGGCCGTTTCGCCTTCCTFGGATATAAATCCG CCG CTCTTCAACGTCG TCTTG CTCTFCGGGAAGAGAAGAGGGCCAAGGAAGTGGCTCGTCTG CAGGAGGGTGAG CAGTG CCAAATGGGTGAGAGGGCCTGTGATGAAG CTCAACGGAGTG CGGATGGAGGG AG CAGTACTGAAGGCAAGAGTCCGG TACTGACCCCCTTGGACGAGAAGAACGTTG CGG CGTTGGGGAGGCATFGAGGTTATTCTFFCCGTAAAGGCAACCAAGAGTAATITAAACCT 11 10 15 20 25 AACGTAGTATTCAGI I I I I I I IGGTTATTAATGAAGFITATGTGCAATAATATTAGTACAT GCCCATCGTATGAAGTGTGTTAGTAAGAA Nucleotide sequence No. 2 cDNA encoding FET3- (ferrous-OZ oxidoreduktas-) component of iron acquisition protein complex [FTR1 / FET3] with high affinity.
    ATGACTAACG CTITG CTCTCTATAG CCGITTTG CTITTCTCGATG CTCTCG CTAG CACAAG CGGAGACGCACACGTTTAATTGGACCACTGGCTGGGACTACAGGAACGTTGATGGGCT AAAGAG CCGTCCCGTGATCACCTGTÅÅTGGCCÅGTICCCÅTGGCCAGATATAACGGTCA ACAAAGGTGACCGTGTG CAGAITFACTTGACCAACGGAATGAACAACACCAATACTTCT CATTTCCACGGTCTCITCCAAAACGGAACCG CCTCTATGGACGGTGTGCCCTTCTTG ATG ACG CG CAATGTCCAATIG CCAGGCAGTACTATG CTTIACAATITCACGGTGGACTACAAT GTAGGCACCTACTG GTACCATTCACACACGGACGGTCAATATGAAGACGGGATGAAAG GTCTITTCATCATCAAGGATGATAG CTTCCCCTACGATTACGATGAGGAACTITCTTTATC GCTTAGTGAGTGGTACCACGACTFGGTCACGGACTTGACGAAGTCGTICATGAGTGTTT ATAATCCGACAGGTG CTGAG CCCATCCCACAGAACTTGATTGTTAACAACACGATGAATC TGACATGGGAAGTCCAGCCCGATACGACGTATCTTTTGAGAATIGTCAACGTGGGTGGG TICGTITCGCAGTACITITGGATCGAGGACCACGAAATGACCGTGGTCGAAATCGACGG TATCACTACCGAGAAGAACGTAACGGATATG CTTTACATCACTGTCG CTCAGAGATATAC AGTCCTGGTTCACACTAAAAACGACACGGACAAAAATTICG CCATCATG CAGAAATITG ATGACACCATGTFGGATGTCATFCCAAGTGATITACAG CTGAATG CAACCTCTTATATGG TCTACAACAAAACCGCTG CTGCCCACACAAAATTACGTGGATFCAATTGATAACTTCT CG TG GA CGATFTCTACTTG CAACCGTACGAGAAAGAAG CCATCTATGGCGAG CCAGATCAT GTGATTACCGTTGACGTTGTFATGGATAACTIGAAAAACGGTGTGAATTACGCCTTCTTC AATAATATCACCTATACTG CACCAAAAGTTCCTACTITGATGACCGTTTTGTCTTCAGGTG ATCÅAG CAAACAACTCCGAAATCTACGGTTCAAACACG CACACTTTCATCCTAGAGAAG GATGAAATCGTGGAGATTGTG CTAAATAACCAGGACACAGGTACCCATCCITFCCATITA CATGGTCACG CTITCCAAACCATCCAGAGAGATCGTACATATGATGATG CCCTAGGTGA AGTTCCTCACAGTFFCGATCCGGACAACCACCCTG CCTCCCAGAATACCCAATGAGAAGA 12 10 15 20 25 GATACITTATACG'ITAGACCACAATCCAATITCGTCATCAGGTAAAGCCGATAACCCAGG TGTTTGGTTCTTCCATTGTCATATCGAATGGCATGTTG CAAGGTFTGGGTCTTGTTCTCGT GGAGGATCCTTTTG GTATCCAAGATG CTCAWCTCAACAACTCAGTGAAAACCACTTAGA AGTTTG CCAGAGTTGCTCTGTGGCCACTGAAGGTAACG CCGCT G CCAATACACTGGATT TAACTGATTTAACTGGTGAAAATGTTCAG CATGCCTTCATTCCTACCGTTWACCAAAAAA G GTATTATTG CCATGACAWCTCCTG CTTTG CCG CAA GTATTCTFGGTATTATCACAATTG TITATGGTATGATGGATATGGAAGATG CGACCGAAAAGGTTATTCGAGACTTG CACGTG GACCCTGAAGTCTTG CTAAATGAGGTTGATGAAAATGAAGAG CGTCAGGTAAACGAAG ATCGTCATTCCACTGAAAAG CATCAATTITAACTAAAGCCAAACGGTTCT FCTAA Nucleotide sequence No. 3 cDNA encoding SUL1, sulfate permease from Saccharomices cerevisiae.
    ATGTCACGTAAGAG CT CGACTG AATATGTG CATAATCAG GAGGATG CTGATATCGAAGT ATFTGAATCAGAATACCG CACATATAGGGAATCTGAGGCGGCAGAAAACAGAGACG GA CTTCACAATGGTGATGAGGAAAATTGGAAGGTTAATAGTAGTAAG CAGAAATTTGGGG TAACGAAAAATGAG CT ATCAGATGTCCTGTACGATTCCATTCCAG CGTATGAAGAGAGC ACAGTCACTTTGAAGGAGTACT ATG ATCATTCTATCAAAAACAATCT AACTG CGAAATCG GCAGGAAGTTACCTCGTATCTCTITITCCTATTATAAAATGGTITCCTCATTATAACTITAC GTGGGGCTATG CTGATITAGTGGCAGGAATTACAGTTGGCTGCGTACTCGTGCCCCAAT CTATGTCATACG CACAAATCG CTAGTTTATCTCCTG AATATGGTTTGTATTCCTCCTFFATT GGTG CGTTTATATATTCWTGTTFG CCACATCGAAAGATGTITGTATTGGTCCGGTCGCTG TAATGTCACTACAAACT G CCAAAGTCATTG CT GAAGTTCTAAAAAAATATCCCGAAGACC AGACAGAAGTTACAG CTCCTATCATTG CAACTACCCTITGTFTG CTITGTGGGATTGTCG CCACTGGGTTGGGTATACTG CGTTTAGGCTFITTAGTGGAACTTATTTCTCTAAATG CT GT TG CTGGCTFCATGACCGGTTCCG CATITAACATCATCTGGGGTCAAATFCCGGCTCTCAT GGGATACAACTCATTAGTGAATACCAGAGAAGCAACGTATAAGGTFGTAATTAACACTC TGAAACATITACCAAACACAAAGTFAGACGCCGTITTGGCTTGATTCCGTTGGTAATCCT CTATGTATGGAAATGGTGGTGTG GTACATTTGGTATAACTTTGGCAGATAGAT ATTATCG 13 10 15 20 25 AAATCAACCAAAG GTAG CAAATAGACTGAAATCCTTCTATTTCTATG CACAAG CTATGAG AAATG CCGTCGTCATAGTAGTTFITACTGCCATATCGTGGAG CATAACAAGAAACAAATC TTCAAAAGACCGTCCAATCAGTATTCTGGGTACAGTTCCCTCGGGCTTAAATGAG GTGG GAGTTATGAAAATCCCAGACGGTCTG CTATCTAATATGAGTTCAGAAATACCTG CTTCAA TTATCGTTCTGGTGTTAGAACACATCG CTATITCAAAATCCTTTG GTAGAATTAACGACTA CAAGGTTGTCCCTGACCAAGAACTTATTG CGATTGGTGTGACAAATTTGATAGGGACATT TITTCACTCATATC CAG CAA CTG GG TCATTITC CAG ATCTG CTITG AAAG CAAAATG TAAC GTG CG CACTCCGTITTCTGGGGTITCACTGGCGGTTG CGTTCTATTAG CACTITATTGTTT AACTGACGCCTTCTFHTCATTCCTAAAGCGACACTATCGGCGGTTATTATTCATG CTGTT TCTGATTTGCTGACTTCTTACAAAACCACCTGGACCTTCTGGAAGACCAACCCGTTAGATT GTATCTCATTTATCGTTACAGTGTTCATCACAGTATTTTCATCCATTGAAAATGGTATATA TITTG CAATGTGTTGGTCATGTG CAATGTTACTATTGAAACAGGCTITCCCTG CTGGTAA ATTCCTTGGTCGTGTTGAGGTGGCAGAAGTATTGAACCCAACAGTACAAGAGGATATTG ATG CTG TG ATATCATCTAATGAATTACCTAATG AA CTG AATAAA CAG GTTAAGTCTACTG TTGAGGTTTTACCAGCCCCAGAGTATAAGTTTAG CGTAAAGTGGGTTCCGTFCGATCATG GATACTCAAGAGAATTGA ATATCAATACCACAGTTCGGCCTCCTCCACCAGGTGTCATAG TCTATCGTITGGGTGATAG CTTTACTTACGTG AACTG CTCAAGGCATTATGACATTATATT TGATCGTATTAAGGAAGAAACAAGGCGAGGCCAACTTATAACCTTAAGGAAAAAGTCA GACCGTCCATGGAATGATCCTGGTGAATGGAAAATGCCAGATTCTITGAAATCACTATTT AAATTFAAACGTCATTCAG CAACAACGAATAGTGACCTACCGATATCGAATGGAAG TAACGGAGAAACATATGAAAAGCCG CTACTGAAAGTCGTCTG CCTGGATTITTCCCAAG CAG TTG CTCAAGTGGATTCAACCG CTGTTCAAAG CCTGGTTGATCTGAGAAAAG CTGTGAAT AG GTATG CG GATAGACAAGTCGAATTCCATTTTGCCG GAATTATATCTCCATGGATCAAA AGAAGTCTTFTGAGTGTTAAATTCGGAACTACAAATGAGGAATATAGTGACGACTCTATT ATCGCTGGCCATTCTAGTTTTCACGTTG CAAAAGTTTTGAAGGATGATGTGGATTATACT GATGAAGACAG CCGTATAAG CACATCTTACAGTAACTATGAAACATTATGTG CTG CAACT GGGACAAATTTACCGTFTTFTCATATCGATATACCCGATITITCTAAATGGGACG1'T'I'AG 14:10 15 20 25 No 4 Nucleotide sequence cDNA encoding the high affinity phosphate transporter PH084.
    ATGAG'ITCCGTCAATAAAGATACTATTCATGTTG CTGAAAGAAGTCTTCATAAAGAACAC CTTACCGAAGGTGGTAACATGGCCTrCCACAACCATTTGAATGATTITG CTCATATrGAAG ATCCTCTGGAAAGAAGAAGATTGGCTTTGGAGTCCATCGATGACGAAGGTTTCGGTTGG CAACAAGTTAAGACCATCTCCATTG CTGGTGTTGGTTTCTTGACAGATrCTTATGATA f T TTG CCATTAATITGGGTATCACTATGATGTCCTACGTTFACTGGCACGGTAGTATGCCAG GTCCAAGTCAAACCTTGTTGAAGGTTTCCACTTCTGTTGGTACTGTTATTG GTCAAT-ITGG TTTTGGTACTTTAGCTGATATTGTTGGTCGTAAGAGAATTTATGGTATGGAACTTATTATC ATGATTGTCTGTACCATTCTG CAAACCACTGTTG CTCATTCTCCTG CTATTAACTrCGTTGC TGTTTTAACATTCTACCGTATrGTCATGGGTATTGGTATCG GTG GTGACTACCCACTATCT TCTATTATTACTTCTGAATTTG CCACTACCAAATGGAGAG GTGCCATCATGGGTG CTGTC TTTG CTAACCAAG CTTGGGGTCAAATCTCCG GTGGTATCATCG CTCTTATCTTGGTTG CT GCTTACAAGGGCGAACTAGAATACG CAAACTCTGGTG CTGAATGTGATG CTAGATGTCA AAAGGCTTGTGACCAAATGTGGAGAATCCTTATTGGGTTGGGTACCGTTCTAGGGTTGG CATGHTGTATTTCAGATTAACTATTCCAGAATCTCCTAGATATCAATTGGATGTTAACGC TAAGTTGGAACTTG CT G CTG CCG CACAAGAACAAGATGGCGAAAAGAAAATTCACGAC ACCAGTGATGAAGACATGGCAATTAACGGTTTGGAAAGAG CTTCTACTGCCGTCG AATC TCTTGACAATCATCCTCCAAAGG CTTCGTTCAAAGATITCT G CAGACATTTTG GTCAATG GAAGTACGGTAAGATITTG CTAGGTACTG CTGGTTCATGGTTTACCTTAGATGTTG CTTT CTACGGGTTGAGTTTAAACAGTG CTGTTATTCTG CAAACCATCGGTTATG CCGGTTCCAA AAACGTITACAAGAAACTGTATGATACTG CTGTCGGTAATCTGATTITGATTTGTG CTGG TTCATTACCTGGTTACTGGGTATCCGTCTTCACTGTCGATATAATCGGTAGAAAACCAATT CAATTAGCCGGTTTCATCATCTTGACCG CHTGTTCTGTGTCATCG GTTTCG CATACCATA AACTTGGTGACCATGGTCTGTTGGCTCTTTACGTCATITGTCAATTCTTCCAAAACTTCGG TCCAAACACAACCACCTTTATTGTTCCTGGTGAGTGTTTCCCAACTCGTTACAGATCTACT 15 10 15 20 25 GCTCATGGTATTTCTG CTG CATCTGGTAAGGTCGGTGCCATTATFG CACAAACCGCTFTG GGTACTCTAATCGACCATAACTGTG CTAGAGACGGTAAGCCAACCAACTGTTG GTTACCT CACGTCATGGAAATTITCG CCTFATFCATGWGTFGGGTATCTTCACAACCTTGTTGATCC CAGAAACTAAGAGAAAGACTCTAGAAGAAATTAACGAG CTATACCACGATGAAATCGAT CCTG CT ACG CTAAACTTCAGAAACAAGAATAATGACATTG AATCTFCCAGCCCATCTCAA CTTCAACATGAAG CATAA BIBLIOGRAPHY first - Julio Alberto Correa, 2008; Siderürgico Process 2.- http://www.cmp.cl 3.- April, E. R. (1974) Introduction to Metallurgy. Marymar Editions. 4.-Gardeweg, M. y Ramirez, C., (1985). Hoja Rio Zapaleri. Carta Geológica de Chile N066, SERNAGEOMIN, Santiago, Chile, 89 pp. 5.- Kulaev and T. Kulakovskaya (2000) POLYPHOSPHATE AND PHOSPHATE PUMP, Annu. Reef. Microbiol. 2000. 54: 709-34. 6.- Manfredo J. Seufferheld, Hector M. Alvarez and Maria E. Farias. (2008), Role of Polyphosphates in Microbial Adaptation to Extreme Environments, APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 5867- 5874. 7.- John W. Mcgrath, John P. Quinn (2000), Intracellular Accumulation of Polyphosphate by the Yeast Candida humicola GI in Response to Acid pH, APPLIED AND ENVIRONMENTAL MICROBIOLOGY 4068- 4073. 8. - Alan Mullan, John W. McGrath, Tom Adamson, Sam lrwin, and John P. Quinn, Pilot-Scale Evaluation of the Application of Low pH-Inducible Polyphosphate Accumulation to the Biological Removal of Phosphate from Wastewaters, Environ. Sci. Technol 40, 296-301. 9.- Buehler, S., A. Oberson, LM. Rao, D.K. Friesen and E. Frossard. (2002) Since much of the soil organic sulfur is present as sulfate esters, aryl and alkylsulfatase enzymes are thought to play a key role in sulfur mineralization. Soil Science Society of America Journal 661868- 877. 16 10 15 20 25 10.— H. Cherest, J. C. Davidian, D. Thomas, V. Benes, W. Ansorge and Y. Surdin-Kerjan (I997). Molecular Characterization of Two High Affinity Sulfate Transporters in Saccharomyces cerevisiae, Genetics 145: 627-635. 11.- Hounayda Mansouri-Bauly, Jörg Kruse, Zuzana Sykorova, Ursula Scheerer, Stanislav Kopriva. (2006) Sulfur uptake in the ectomycorrhizal fungus Laccaria bicolor S238N. Mycorrhiza 16: 421-427. 12.- Hatem Rouached, David Secco, A. Bulak Arpat (2009) Getting the mos tsulfate from soil: Regulation of sulfate uptake transporters in Arabidopsis. Journal of Plant Physiology 166 893- 902. 13.- George A. Marzluf (l997) Molecular genetics of sulfur assimilation in filamentous fungi and yeast.
    Annu. Reef. Microbiol. 51: 73- 96. 14.- Sebastian Pilsyk and Andrzej Paszewski (2009) Sulfate permeases - phylogenetic diversity of sulfate transport. Acta bioquimica polonia 56: 375-384. 15.-Marianne Wessling-Resnick (1999) Biochemistry of Iron Uptake. Critical Reviews in Biochemistry and Molecular Biology, 34 (5): 285-314. 16.-Wissnicka R, Krzepilko A, Wawryn J, Bilinski T. (1997) Iron toxicity in yeast. Acta Microbiol Po | .; 46 (4): 339-47. 17.- Anthony Van Ho, Diane McVeyWard, y Jerry Kaplan. (2002) TRANSITION METAL TRANSPORT IN YEAST. Annu. Reef. Microbiol .. 56: 237-61. 18.- Ernest Kwok and Daniel Kosman, (2005), Iron in yeast: Mechanisms involved in homeostasis, Vol. 14360-99. 19.- Dennis D. Wykoff and Erin K. 0'Shea (2001). Genetics 159: 1491 -1499. 17
    权利要求:
    Claims (22)
    [1]
    A recombinant cell whose genes encoding the high affinity iron uptake protein complex [FTR1 / FET3] have been mutated artificially, while this cell has been transformed with two exogenous genes encoding PHO84 and SUL1, both from Saccharomyces cerevisiae, which can be used to remove phosphorus and sulfur from concentrate and / or iron ore, CHARACTERIZED in that said mutated and double-transformed cell is tolerant to high iron conditions (over 50% by weight) due to the lack of complex [FTR1 / FET3] (nucleotide sequences no. on the other hand, due to expression of nucleotide sequences encoding SUL1 and PHO84 from Saccharomyces cerevisiae (nucleotide sequences Nos. 3 and 4), the same cell can take up and accumulate phosphorus and / or sulfur from culture medium.
    [2]
    Cell according to claim 1, CHARACTERIZED in that it is a eukaryotic cell.
    [3]
    Cell according to claim 1, CHARACTERIZED in that it is a yeast fungus.
    [4]
    Cell according to claim / CHARACTERIZED in that it is a bacterium.
    [5]
    Use of the transformed cell according to any one of claims 1-4, CHARACTERIZED in that said cell can be used to remove phosphorus and / or sulfur from liquid or solid medium which is iron concentrated or comes from iron ore, which includes the incorporation of said cell in said iron-concentrated medium, wherein the mutated and double-transformed cell proliferates while gradually incorporating and accumulating phosphorus and / or sulfur in its biomass, thus removing them from the medium.
    [6]
    Use of the cell according to claim 5, CHARACTERIZED in that said cell can be used to remove phosphorus and / or sulfur from iron-concentrated media derived from industrial processes. 18 10 15 20 25 30 35
    [7]
    Use of the cell according to claim 6, CHARACTERIZED in that said cell can be used to remove phosphorus and / or sulfur from aqueous media contaminated with phosphorus and / or sulfur resulting from extraction processes.
    [8]
    Use of the cell according to claim 5, CHARACTERIZED in that said cell can be used to remove phosphorus and / or sulfur from iron ore resulting from extraction processes.
    [9]
    Use of the cell according to claim 7, CHARACTERIZED in that said cell can be used to remove phosphorus and / or sulfur from media contaminated with sulfur and phosphorus resulting from extraction processes.
    [10]
    Use of the cell according to any one of claims 5, 6 or 8, CHARACTERIZED in that said cell can be used to remove phosphorus from iron-concentrated media.
    [11]
    Use of the cell according to any one of claims 5, 6 or 8, CHARACTERIZED in that said cell can be used to remove sulfur from iron-concentrated media.
    [12]
    Use of the cell according to any one of claims 7 or 9, CHARACTERIZED in that said cell can be used to decontaminate liquid or solid media contaminated with phosphorus.
    [13]
    Use of the cell according to any one of claims 7 or 9, CHARACTERIZED in that said cell can be used to decontaminate liquid or solid media contaminated with sulfur.
    [14]
    Use of the cell according to any one of claims 1-4, CHARACTERIZED in that it can be used to recover phosphorus and / or sulfur from an iron-concentrated medium, which comprises incorporating said cell into said iron-concentrated medium, where the mutated and double-transformed cell multiplies while gradually incorporating and accumulating phosphorus and / or sulfur in its biomass, thus recovering both substances from the medium, enabling a subsequent recovery of phosphorus and / or sulfur from the biomass of the cell.
    [15]
    Use of the cell according to claim 14, CHARACTERIZED in that said cell can be used to recover phosphorus.
    [16]
    Use of the cell according to claim 14, CHARACTERIZED in that said cell can be used to recover sulfur.
    [17]
    Process for removing phosphorus and / or sulfur from a liquid or solid body, CHARACTERIZED in that it comprises the steps of a) placing said medium to be emptied of phosphorus and sulfur at a temperature between -20 ° C and 40 ° C, b) incorporating the transformed cell according to any one of claims 1-4 into the medium to be emptied as described in step a), c) enabling propagation of said cell in said medium, until the cell density reaches an OD value (600 nm) of 0 , 75 to 1, and d) remove the cells by sedimentation or filtration.
    [18]
    A method according to claim 17, CHARACTERIZED in that it enables the removal of phosphorus from a medium.
    [19]
    A method according to claim 17, CHARACTERIZED in that it enables the removal of sulfur from a medium.
    [20]
    Process for recovering phosphorus and / or sulfur from a medium, CHARACTERIZED in that it comprises the steps of a) placing said medium to be emptied at a temperature between -20 ° C and 40 ° C, b) incorporating it mutated and double transformed the cell according to any one of claims 1-4 in the medium to be emptied as described in step a), c) enabling propagation of said cell in said medium, until the cell density reaches an OD value (600 nm) of 0.75 to 1 , d) collecting the cells by sedimentation or filtration, e) lysing the cells and extracting the phosphorus and / or sulfur by chromatography.
    [21]
    A method according to claim 20, CHARACTERIZED in that it enables the recovery of phosphorus from a medium.
    [22]
    Process according to Claim 20, CHARACTERIZED in that it enables the recovery of sulfur from a medium. 20
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    同族专利:
    公开号 | 公开日
    CL2011000609A1|2012-02-10|
    WO2012126134A2|2012-09-27|
    AU2012231635A1|2013-11-07|
    BR112013024068A2|2017-10-17|
    PE20140929A1|2014-08-24|
    引用文献:
    公开号 | 申请日 | 公开日 | 申请人 | 专利标题
    法律状态:
    2015-03-31| NAV| Patent application has lapsed|
    优先权:
    申请号 | 申请日 | 专利标题
    CL2011000609A|CL2011000609A1|2011-03-22|2011-03-22|Recombinant cell comprising: the endogenous ftr1 / fet3 complex removed and inserted the coding sequences for a sulfur permeaseand a high affinity phosphate transporter , useful for extracting phosphorus and / or sulfur from a medium concentrated in iron.|
    PCT/CL2012/000014|WO2012126134A2|2011-03-22|2012-03-22|Recombinant cell that has a mutated cellular iron-transport mechanism and is transformed with a sequence that codes for a sulphur permease and with a sequence that codes for a high-affinity phosphate transporter, suitable for extracting phosphorus and/or sulphur from a solid or liquid medium that may have a high iron content|
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