Fast quantitative detection method for methanogenic archaea in oil-water well output liquid
A quantitative detection method and a technology of methanogenic archaea, which are applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of poor detection accuracy, long cultivation time, missed detection of methanogenic bacteria, etc., and achieve sensitivity and High accuracy, short detection time and high reproducibility results
Inactive Publication Date: 2014-11-19
CHINA PETROLEUM & CHEM CORP +1
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Problems solved by technology
[0003] Quantitative detection of methanogenic archaea in oil and water wells can better analyze the mechanism of endogenous microbial oil displacement. At present, the detection methods for methanogenic archaea mainly use the maximum likelihood number method and direct microscopy. The cultivation takes a long time, and bacterial cultivation usually takes 1 to 3 months. After the cultivation, the methane gas generated needs to be detected by gas chromatography. This process is time-consuming and labor-intensive; Simulating the reservoir environment will cause some methanogenic bacteria with different physiological characteristics to be missed, and the detection accuracy is poor; for direct microscopy, because it cannot effectively and accurately distinguish methanogenic archaea from other bacteria in the mixed flora, Therefore, this method can only be used for quantitative detection of some purely cultured methanogens, and is not suitable for the detection of oil reservoir environmental samples.
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Embodiment 1
[0059] Utilize the method of the present invention to the detection of methanogenic archaea in the output fluid samples of 5 oil wells and 1 water well in a certain block of Shengli Oilfield, and its specific steps are:
[0060] 1. Synthesis of specific primers
[0061] Specific primer pair sequence of the present invention:
[0062] Mcr A-R: 5`-CGTTCATBGCGTAGTTVGGRTAGT-3`
[0063] Mcr A-F: 5`-GGTGGTGTMGGDTTCACMCARTA-3`
[0064] 2. Preparation of standard plasmids
[0065] The preparation method of the standard plasmid is as follows:
[0066] (1) Cultivation of methanogenic bacteria
[0067] The DNA of a strain of methanogenic archaea (Methane mane basilicate) was used to construct the standard plasmid of the methyl-CoA reductase gene fragment, and the formula of 1 L of methanogenic archaea medium was: NH 4 Cl 1g / L, K 2 HPO 4 0.3g / L, KH 2 PO 4 0.3g / L, KCl 1g, CaCl 2 0.1g / L, MgCl 2 0.2g / L, NaCl 5g / L, MgSO 4 0.2g / L, yeast powder 2.5g / L, peptone 0.5g / L, trace elements...
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The invention discloses a fast quantitative detection method for methanogenic archaea in oil-water well output liquid. The fast quantitative detection method comprises the following step: 1, compounding specific primers; 2, preparing standard quality grains; 3, establishing a fluorogenic quantitative PCR reaction process and a reaction system; 4, drawing a standard curve; 3, evaluating the method; 6, sampling a field sample, collecting thalli, and extracting DNA; 7, quantitatively detecting methanogenic archaea of the field sample. The method is high in accuracy, strong in specificity, simple in operation, high in repeatability and short in detection time; the process from extraction of sample DNA to the obtaining of the detection result requires less than 3 h; high performance detection can be performed to 96 samples at the same time; the method is used for fast quantitative detection of methanogenic archaea in oil-water well output liquid.
Description
technical field [0001] The invention belongs to the technical field of tertiary oil recovery, and in particular relates to a quantitative detection method for methanogenic archaea in the output fluid of oil-water wells. Background technique [0002] Microbial oil recovery is an emerging tertiary oil recovery technology, through the combined action of endogenous or exogenous injected microorganisms and their metabolites to increase crude oil production. Unlike chemical flooding, this technology has the advantages of low cost, wide adaptability, simple operation, and environmental friendliness, so this technology is of great significance to the further development of my country's petroleum industry. Endogenous microbial flooding activates different functional microorganisms in the oil reservoir by injecting suitable nutrients into the formation, such as hydrocarbonophilic bacteria, bacteria producing biosurfactants, and some methanogenic archaea, among which the methanogenic a...
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Login to View More IPC IPC(8): C12Q1/68C12Q1/06
CPCC12Q1/6851C12Q2531/113C12Q2563/107C12Q2545/113
Inventor张绍东胡婧曹功泽王静宋欣段传慧杜春安徐鹏王刚徐闯宋永亭
OwnerCHINA PETROLEUM & CHEM CORP