Method for determining a metal present in a sample
First Claim
1. A method for determining a heavy metal in a sample comprising;
- a) providing a cell comprising a recombinant DNA plasmid, said plasmid comprising a gene coding for an insect luciferase or a green fluorescent protein (GFP) marker protein, wherein the copy number of said plasmid is under the control of a promoter regulatable by a heavy metal, and wherein said promoter controls the origin of replication of said plasmid;
b) contacting said cell with a sample which contains said heavy metal;
c) allowing said heavy metal to affect said cell, whereafter the amount of said marker protein is determined;
d) comparing the amount of said marker protein with a control in which no metal was present or in which the heavy metal was present in a known amount, whereby the presence and/or amount of said heavy metal is determined.
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Abstract
The invention relates to a method for determining a metal present in a sample, in which method
a) a recombinant DNA plasmid the copy number of which is under the control of a promoter regulatable by the metal is transferred into the cell, or a recombinant DNA plasmid the copy number of which can be varied between 1 and 2000/cell is transferred into the cell, and the plasmid contains as a marker protein a DNA sequence coding for a virus, procaryotic cell or eucaryotic cell protein or for a portion thereof essential for its biological activity, the expression of the DNA sequence being under the control of a promoter regulatable by the metal and being controlled by negative and/or positive feedback;
b) the cell containing the recombinant DNA plasmid is contacted with a metal-containing sample;
c) the metal is allowed to affect for a suitable time the cell which contains the recombinant DNA plasmid, whereafter the amount of recombinant DNA plasmid or the amount of protein encoded by it is determined by either physical or chemical means;
d) the amount of recombinant DNA plasmid or the protein encoded by it is compared by chemical or physical means to a control test in which metal was not present and/or it was present in a known amount in a reaction, whereby the presence and/or amount of the metal can be determined.
20 Citations
34 Claims
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1. A method for determining a heavy metal in a sample comprising;
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a) providing a cell comprising a recombinant DNA plasmid, said plasmid comprising a gene coding for an insect luciferase or a green fluorescent protein (GFP) marker protein, wherein the copy number of said plasmid is under the control of a promoter regulatable by a heavy metal, and wherein said promoter controls the origin of replication of said plasmid; b) contacting said cell with a sample which contains said heavy metal; c) allowing said heavy metal to affect said cell, whereafter the amount of said marker protein is determined; d) comparing the amount of said marker protein with a control in which no metal was present or in which the heavy metal was present in a known amount, whereby the presence and/or amount of said heavy metal is determined. - View Dependent Claims (2, 3, 4, 5, 6, 26, 27, 28, 29, 30, 31, 32, 34)
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7. A method for the determination of a heavy metal in a sample, comprising:
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a) providing a cell comprising a recombinant DNA plasmid, said plasmid comprising a gene coding for an insect luciferase or a green fluorescent protein (GFP) marker protein, and having a copy number between 1 and 2000/cell, wherein the expression of said marker protein sequence being a heavy metal, and wherein said promoter controls the origin of the replication of said plasmid; b) contacting said cell with a sample which contains said heavy metal; c) allowing said heavy metal to affect said cell, whereafter the amount of said marker protein is determined; d) comparing the amount of said marker protein with a control in which no metal was present or in which it was present in a known amount, whereby the presence and amount of said heavy metal is determined. - View Dependent Claims (8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 33)
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22. The plasmid pMV1 (DSM Deposit No. 8708).
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23. The plasmid pMV3 (DSM Deposit No. 8893).
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24. The plasmid pTOO21 (DSM Deposit No. 9666).
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25. The plasmid pTOO23 (DSM Deposit No. 9667).
Specification