CN102128938B - Anabolic steroids and polycyclic aromatic hydrocarbon high-efficiency bioluminescence sensor and construction method thereof - Google Patents
Anabolic steroids and polycyclic aromatic hydrocarbon high-efficiency bioluminescence sensor and construction method thereof Download PDFInfo
- Publication number
- CN102128938B CN102128938B CN 201010595059 CN201010595059A CN102128938B CN 102128938 B CN102128938 B CN 102128938B CN 201010595059 CN201010595059 CN 201010595059 CN 201010595059 A CN201010595059 A CN 201010595059A CN 102128938 B CN102128938 B CN 102128938B
- Authority
- CN
- China
- Prior art keywords
- plasmid
- gfp3
- centrifugal
- sequence
- supernatant
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Images
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
Abstract
The invention relates to an anabolic steroids and polycyclic aromatic hydrocarbon high-efficiency bioluminescence sensor, belonging to the technical field of molecular biology; the bioluminescence sensor is characterized in that plasmids pK-3[alpha]-GFP3 and p6 are respectively transformed into colibacillus and respectively prepared into cell-free system testing agent; 3[alpha]-HSD promoters such as regulatory sequence 486 base pair and GFP reporter gene 722 base pair are inserted at the down stream of the cis-[belta]-galactosidase (LacZ) promoter gene of recombinant plasmid pK-3[alpha]-GFP3 in sequence; the regulatory sequence of 3[alpha]-HSD on p6 plasmid can be induced to express protein with regulatory function; the protein can promote the promoter on the plasmids pK-3[alpha]-GFP3 so as to express the GFP protein; at the moment, the expression quantity of the GFP is controlled by the quantity of induction substrate strictly. The bioluminescence sensor and the construction method thereof have the following benefits: aiming at the shortcomings of strict instrument requirement, long detection cycle, high cost, low sensitivity and single test piece in the existing testing system, the bioluminescence sensor and the construction method thereof have wide test linearity width, simple technical operation and low instrument requirement, and are applicable to popularize and apply to detect anabolic steroids and polycyclic aromatic hydrocarbon pollutant in the environment and food.
Description
Technical field
The invention belongs to technical field of molecular biology.
Background technology
In recent years, natural, artificial synthetic steroid and polycyclic aromatic hydrocarbon compounds is unconfined is discharged in physical environment, the hormonal activity of this pollutant and to the negative influence of biosome extremely people pay close attention to.Because it is widely distributed, all have hormonal activity under extremely low concentration, environment and human health are produced serious harm, so can new technology sensitive, that detect fast and efficiently steroid and polycyclic aromatic hydrocarbon compounds demand development urgently.Traditional detection analytical technology is mainly gas chromatography-mass spectrum (GC-MS), liquid chromatography-mass spectrography (HC-MS) at present, and domestic and international expert has been developed into multiple steroid hormone detection system based on the research of human body steroid receptor in addition.These detection system great majority need human body or yeast cells to cultivate, and sense cycle is long, cost is high, exist in addition sensitivity low, can only detect the drawback such as a kind of compound, it is extremely limited therefore these technology being applied to the environment measuring aspect.
Comamonas testosteroni (C.testosteroni, C.T) remarkable characteristic is that it has specificity degradation of steroid class and polycyclic aromatic hydrocarbon compounds enzyme gene, and existing research about its degradation of steroid class and polycyclic aromatic hydrocarbon compounds gene expression and steroid hormone and polycyclic aromatic hydrocarbon compounds acknowledgement mechanism.Whether can be with C.T take steroid and polycyclic aromatic hydrocarbon compounds as signal scheme, build a kind of sensitivity, fast, simple biology sensor, be used for steroid and polycyclic aromatic hydrocarbon compounds potpourri in testing environment.
Summary of the invention
The objective of the invention is: a kind of steroid hormone and palycyclic aromatic high-performance bio fluorescent optical sensor and construction method are provided, plasmid pK-3 α-GFP3 and p6 are transformed respectively in Escherichia coli, set up a kind of bioluminescence detection system of Green Fluorescent Protein, detected object is steroid hormone and the large class material of palycyclic aromatic two.
Technical scheme of the present invention is:
The present invention has built based on the steroid hormone of Comamonas testosteroni (C.testosteroni, C.T) and palycyclic aromatic bioluminescence detection system.Comamonas testosteroni can be with steroid hormones such as stosterones as sole carbon source and the energy, the materials such as palycyclic aromatic of the non-steroid of can also degrading, and (metabolism of the enzyme such as 3 α-HSD) digests this class substrate by 3 α-hydroxysteroid dehydrogenase.Green fluorescent protein (GFP) is the reporter gene that is widely used in recent years the biological marker field, the protein that its gene produces, under the light of blue wavelength region excites, green fluorescence can be sent, the relative content of specific substrate can be calculated according to its fluorescence intensity by the reference standard curve.
The present invention transforms plasmid pK-3 α-GFP3 and p6 respectively in Escherichia coli and is prepared into respectively acellular system detection reagent, has set up a kind of bioluminescence detection system of Green Fluorescent Protein.3 α-regulating and controlling sequence (seeing sequence 2) 470 base-pairs and GFP reporter gene (seeing sequence 3) 717 base-pairs such as HSD promoter have been inserted in the beta galactosidase of taking advantage of a situation (LacZ) promoter gene (seeing sequence 1) downstream at recombinant plasmid pK-3 α-GFP3 successively; Recombinant plasmid p6 contains the whole controlling genes of 3 α-HSD (seeing sequence 4), totally 5257 base-pairs.When having the substrates such as estrogen in environment, can induce the albumen of the regulating and controlling sequence expression regulation effect of 3 α on the p6 plasmid-HSD, this albumen can make the promoter on plasmid pK-3 α-GFP3 start and then give expression to GFP albumen, and the expression of GFP strictly is subject to inducing the control of amount of substrate at this moment.
The LacZ promoter gene of taking advantage of a situation in plasmid pK-3 α-GFP3 can act on regulating and controlling sequence and the promoter gene of 3 α in itself plasmid-HSD, promotes green fluorescent protein great expression thereafter, and fluorescence signal is had amplification, humidification.According to the fluorescence intensity size of this fluorescent optical sensor, can calculate the relative content of specific substrate by the reference standard curve.
Construction method of the present invention is:
(1) plasmid pK-3 α-GFP3 is transformed in Escherichia coli, cultivates 12 to 14 hours, and after cultivating, Escherichia coli were inoculated in nutrient culture media in 1: 20 by volume, enlarge and cultivate, after cultivating, bacterium liquid is centrifugal, abandons supernatant, add sterilized water washing thalline, the centrifugal supernatant of abandoning, repeated washing twice adds sterilized water and lysozyme, resuspended thalline,-20 ℃ to 25 ℃ multigelations three times, centrifugal, get the acellular system cytoplasm (can preserve month for 20 ℃) that supernatant is preparation.
(2) plasmid p6 is transformed in Escherichia coli, cultivated 12 to 14 hours, after cultivating, Escherichia coli were inoculated in nutrient culture media in 1: 20 by volume, enlarge and cultivate, after cultivating, bacterium liquid is centrifugal, abandons supernatant, add sterilized water washing thalline, the centrifugal supernatant of abandoning, repeated washing twice adds sterilized water and lysozyme, resuspended thalline,-20 ℃ to 25 ℃ multigelations three times, centrifugal, get the acellular system cytoplasm (can preserve month for 20 ℃) that supernatant is preparation.
(3) the acellular system cytoplasm of the acellular system cytoplasm of plasmid pK-3 α-GFP3 and plasmid P6 measured respectively protein content and be prepared into respectively the working fluid of protein content=0.1mg/mL, two kinds of working fluids mix by volume at 1～2: 1～10.Use this and mix acellular system construction high-performance bio fluorescent optical sensor.
Set up bioluminescence standard detection curve
(1) with detection substrate standard items anhydrous alcohol solution, compound concentration is 10 respectively
-3M, 10
-6M, 10
-9M, 10
-12M, 10
-15The standard detection liquid of M/L.
(2) get acellular system detection reagent and add respectively standard detection liquid, blank adds absolute ethyl alcohol, and is standing, detects its fluorescence radiation value, Criterion detection curve with fluorescence microplate reader.
Detected object of the present invention is:
Steroid hormone and palycyclic aromatic two class materials, the sample detection concentration range is: 10
-15To 10
-3Mole every liter.
The invention has the beneficial effects as follows:
Instrument for existing detection system is strict, sense cycle is long, cost is high, sensitivity is low, detect the drawbacks such as thing is single, the present invention detects linear wide ranges, technical operation is simple, instrument is less demanding, suitablely is applied to steroid hormone and polycyclic aromatic hydrocarbon pollutant in testing environment, food.
Description of drawings
Fig. 1 is plasmid pK-3 α-GFP3 schematic diagram;
Fig. 2 is plasmid p6 schematic diagram;
Fig. 3 is stosterone fluoroscopic examination figure as a result;
Fig. 4 is estradiol fluoroscopic examination figure as a result;
Fig. 5 is fluorescent naphthalimide testing result figure.
Embodiment
Embodiment 1 sets up bioluminescence examination criteria curve as an example of stosterone example
1, the cytoplasmic preparation of acellular system:
(1) plasmid pK-3 α-GFP3 is transformed in Escherichia coli, 37 ℃, cultivated 12 to 14 hours in 180～200 rpms of constant-temperature shaking incubators, be inoculated in the Escherichia coli after cultivating in the SIN nutrient culture media in 1: 20 by volume, 37 ℃, be cultured to OD595=3.0 in 180～200 rpms of constant-temperature shaking incubators, after cultivating, 4000 rpms of thalline were abandoned supernatant in centrifugal 20～30 minutes, add sterilized water washing thalline, 4000 rpms centrifugal 20～30 minutes, abandon supernatant, twice of repeated washing, adding sterilized water and final concentration is 100 μ g/mL lysozymes, resuspended thalline,-20 ℃ to 25 ℃ multigelations three times, 10000 rpms centrifugal 20～30 minutes, get supernatant and be the acellular system cytoplasm (can preserve month for 20 ℃) of preparation.
(2) plasmid p6 is transformed in Escherichia coli, 37 ℃, cultivated 12 to 14 hours in 180～200 rpms of constant-temperature shaking incubators, be inoculated in the Escherichia coli after cultivating in the SIN nutrient culture media in 1: 20 by volume, 37 ℃, be cultured to OD in 180～200 rpms of constant-temperature shaking incubators
595=3.0, after cultivating, 4000 rpms of thalline were abandoned supernatant in centrifugal 20～30 minutes, add sterilized water washing thalline, 4000 rpms centrifugal 20～30 minutes, abandon supernatant, twice of repeated washing, adding sterilized water and final concentration is 100 μ g/mL lysozymes, resuspended thalline ,-20 ℃ to 25 ℃ multigelations three times, 10000 rpms centrifugal 20～30 minutes, get the acellular system cytoplasm (can preserve month for 20 ℃) that supernatant is preparation.
(3) the acellular system cytoplasm of the acellular system cytoplasm of plasmid pK-3 α-GFP3 and plasmid P6 measured respectively protein content and be prepared into respectively the working fluid of protein content=0.1mg/mL, and mixing by 1: 1 by volume.
2, set up bioluminescence standard detection curve
(1) with stosterone standard items anhydrous alcohol solution, compound concentration is the standard detection liquid of 1mM/L, 1 μ M/L, 1nM/L, 1pM/L, 1fM/L respectively.
(2) get 100 μ L mixing acellular system cytoplasms and add respectively 2 μ L standard detection liquid, blank adds 2 μ L absolute ethyl alcohols, 30 ℃ after standing 10 minutes, detects its fluorescence radiation value, Criterion detection curve with fluorescence microplate reader.
Fluoroscopic examination result such as following table:
Table 1. stosterone fluoroscopic examination result
The stosterone detectability is respectively 0-1 μ M/L, and fluorescence intensity and concentration meet cubic polynomial curve: y=-284.24x
3+ 2225.9x
2-3172.7x+38422, R
2=0.994.
Embodiment 2 sets up bioluminescence standard detection curve as an example of estradiol example
1, the cytoplasmic preparation of acellular system:
(1) plasmid pK-3 α-GFP3 is transformed in Escherichia coli, 37 ℃, cultivated 12 to 14 hours in 180～200 rpms of constant-temperature shaking incubators, be inoculated in the Escherichia coli after cultivating in the SIN nutrient culture media in 1: 20 by volume, 37 ℃, be cultured to OD in 180～200 rpms of constant-temperature shaking incubators
595=3.0, after cultivating, 4000 rpms of thalline were abandoned supernatant in centrifugal 20～30 minutes, add sterilized water washing thalline, 4000 rpms centrifugal 20～30 minutes, abandon supernatant, twice of repeated washing, adding sterilized water and final concentration is 100 μ g/mL lysozymes, resuspended thalline ,-20 ℃ to 25 ℃ multigelations three times, 10000 rpms centrifugal 20～30 minutes, get the acellular system cytoplasm (can preserve month for 20 ℃) that supernatant is preparation.
(2) plasmid p6 is transformed in Escherichia coli, 37 ℃, cultivated 12 to 14 hours in 180～200 rpms of constant-temperature shaking incubators, be inoculated in the Escherichia coli after cultivating in the SIN nutrient culture media in 1: 20 by volume, 37 ℃, be cultured to OD in 180～200 rpms of constant-temperature shaking incubators
595=3.0, after cultivating, 4000 rpms of thalline were abandoned supernatant in centrifugal 20～30 minutes, add sterilized water washing thalline, 4000 rpms centrifugal 20～30 minutes, abandon supernatant, twice of repeated washing, adding sterilized water and final concentration is 100 μ g/mL lysozymes, resuspended thalline ,-20 ℃ to 25 ℃ multigelations three times, 10000 rpms centrifugal 20～30 minutes, get the acellular system cytoplasm (can preserve month for 20 ℃) that supernatant is preparation.
(3) the acellular system cytoplasm of the acellular system cytoplasm of plasmid pK-3 α-GFP3 and plasmid P6 measured respectively protein content and be prepared into respectively the working fluid of protein content=0.1mg/mL, and mixing by 1: 1 by volume.
2, set up bioluminescence standard detection curve
(1) with estradiol standard items anhydrous alcohol solution, compound concentration is the standard detection liquid of 1mM/L, 1 μ M/L, 1nM/L, 1pM/L, 1fM/L respectively.
(2) get 100 μ L mixing acellular system cytoplasms and add respectively 2 μ L standard detection liquid, blank adds 2 μ L absolute ethyl alcohols, 30 ℃ after standing 10 minutes, detects its fluorescence radiation value, Criterion detection curve with fluorescence microplate reader.
Fluoroscopic examination result such as following table:
Table 2. estradiol fluoroscopic examination result
The estradiol detectability is respectively 0-1 μ M/L, and fluorescence intensity and concentration meet cubic polynomial curve: y=3.1852x
3-344.84x
2+ 3185.1x+33801, R
2=0.9969.
Embodiment 3 sets up bioluminescence standard detection curve as an example of naphthalene example
1, the cytoplasmic preparation of acellular system:
(1) plasmid pK-3 α-GFP3 is transformed in Escherichia coli, 37 ℃, cultivated 12 to 14 hours in 180～200 rpms of constant-temperature shaking incubators, be inoculated in the Escherichia coli after cultivating in the SIN nutrient culture media in 1: 20 by volume, 37 ℃, be cultured to OD in 180～200 rpms of constant-temperature shaking incubators
595=3.0, after cultivating, 4000 rpms of thalline were abandoned supernatant in centrifugal 20～30 minutes, add sterilized water washing thalline, 4000 rpms centrifugal 20～30 minutes, abandon supernatant, twice of repeated washing, adding sterilized water and final concentration is 100 μ g/mL lysozymes, resuspended thalline ,-20 ℃ to 25 ℃ multigelations three times, 10000 rpms centrifugal 20～30 minutes, get the acellular system cytoplasm (can preserve month for 20 ℃) that supernatant is preparation.
(2) plasmid p6 is transformed in Escherichia coli, 37 ℃, cultivated 12 to 14 hours in 180～200 rpms of constant-temperature shaking incubators, be inoculated in the Escherichia coli after cultivating in the SIN nutrient culture media in 1: 20 by volume, 37 ℃, be cultured to OD in 180～200 rpms of constant-temperature shaking incubators
595=3.0, after cultivating, 4000 rpms of thalline were abandoned supernatant in centrifugal 20～30 minutes, add sterilized water washing thalline, 4000 rpms centrifugal 20～30 minutes, abandon supernatant, twice of repeated washing, adding sterilized water and final concentration is 100 μ g/mL lysozymes, resuspended thalline ,-20 ℃ to 25 ℃ multigelations three times, 10000 rpms centrifugal 20～30 minutes, get the acellular system cytoplasm (can preserve month for 20 ℃) that supernatant is preparation.
(3) the acellular system cytoplasm of the acellular system cytoplasm of plasmid pK-3 α-GFP3 and plasmid P6 measured respectively protein content and be prepared into respectively the working fluid of protein content=0.1mg/mL, and mixing by 1: 1 by volume.
2, set up bioluminescence standard detection curve
(1) with naphthalene standard items anhydrous alcohol solution, compound concentration is the standard detection liquid of 1mM/L, 1 μ M/L, 1nM/L, 1pM/L, 1fM/L respectively.
(2) get 100 μ L mixing acellular system cytoplasms and add respectively 2 μ L standard detection liquid, blank adds 2 μ L absolute ethyl alcohols, 30 ℃ after standing 10 minutes, detects its fluorescence radiation value, Criterion detection curve with fluorescence microplate reader.Fluoroscopic examination result such as following table:
Table 3. fluorescent naphthalimide testing result
The estradiol detectability is respectively 0-1 μ M/L, and fluorescence intensity and concentration meet cubic polynomial curve: y=12.815x
3-125.23x
2+ 1444.4x+36335, R
2=0.9298.
Claims (5)
1. a steroid hormone and palycyclic aromatic high-performance bio fluorescent optical sensor, it is characterized in that: plasmid pK-3 α-GFP3 and p6 are transformed respectively in Escherichia coli and be prepared into respectively acellular system detection reagent, set up a kind of bioluminescence detection system of Green Fluorescent Protein; Be that to have inserted successively 3 α-HSD promoter regulation sequence be that 470 base-pairs and the GFP albumen reporter gene of sequence 2 is 717 base-pairs of sequence 3 for sequence 1 downstream at the beta galactosidase promoter gene of taking advantage of a situation of recombinant plasmid pK-3 α-GFP3; It is sequence 4 that recombinant plasmid p6 contains the whole controlling genes of 3 α-HSD, totally 5257 base-pairs; When having the estrogen substrate in environment, can induce the albumen of the regulating and controlling sequence expression regulation effect of 3 α on the p6 plasmid-HSD, this albumen can make the promoter on plasmid pK-3 α-GFP3 start and then give expression to GFP albumen, and this moment, GFP protein expression amount strictly was subject to inducing the control of amount of substrate.
2. the construction method of a steroid hormone and palycyclic aromatic high-performance bio fluorescent optical sensor, its construction method is:
A, plasmid pK-3 α-GFP3 are transformed in Escherichia coli, cultivate 12 to 14 hours, and after cultivating, Escherichia coli were inoculated in nutrient culture media in 1: 20 by volume, enlarge and cultivate, after cultivating, bacterium liquid is centrifugal, abandons supernatant, add sterilized water washing thalline, the centrifugal supernatant of abandoning, repeated washing twice adds sterilized water and lysozyme, resuspended thalline,-20 ℃ to 25 ℃ multigelations three times, centrifugal, get the acellular system cytoplasm that supernatant is preparation;
B, plasmid p6 is transformed in Escherichia coli, cultivated 12 to 14 hours, after cultivating, Escherichia coli were inoculated in nutrient culture media in 1: 20 by volume, enlarge and cultivate, after cultivating, bacterium liquid is centrifugal, abandons supernatant, add sterilized water washing thalline, the centrifugal supernatant of abandoning, repeated washing twice adds sterilized water and lysozyme, resuspended thalline,-20 ℃ to 25 ℃ multigelations three times, centrifugal, get the acellular system cytoplasm that supernatant is preparation;
C, the acellular system cytoplasm of the acellular system cytoplasm of plasmid pK-3 α-GFP3 and plasmid P6 is measured respectively protein content and be prepared into respectively the working fluid of protein content=0.1mg/mL, two kinds of working fluids mix by volume at 1～2: 1～10; Use this and mix acellular system construction high-performance bio fluorescent optical sensor;
D, with detection substrate standard items anhydrous alcohol solution, compound concentration is 10 respectively
-3M, 10
-6M, 10
-9M, 10
-12M, 10
-15The standard detection liquid of M/L;
F, get the acellular system and detect reagent and add respectively standard detection liquid, blank adds absolute ethyl alcohol, and is standing, detects its fluorescence radiation value, Criterion detection curve with fluorescence microplate reader.
3. a kind of steroid hormone according to claim 1 and palycyclic aromatic high-performance bio fluorescent optical sensor, it is characterized in that: the LacZ promoter gene of taking advantage of a situation in plasmid pK-3 α-GFP3 can act on regulating and controlling sequence and the promoter gene of 3 α in itself plasmid-HSD, green fluorescent protein great expression after promotion plasmid pK-3 α-GFP3 has amplification, humidification to fluorescence signal.
4. a kind of steroid hormone according to claim 1 and palycyclic aromatic high-performance bio fluorescent optical sensor, is characterized in that: according to the fluorescence intensity size of this fluorescent optical sensor, calculate the relative content of specific substrate by the reference standard opisometer.
5. a kind of steroid hormone according to claim 1 and palycyclic aromatic high-performance bio fluorescent optical sensor, it is characterized in that: detected object is steroid hormone and palycyclic aromatic two class materials, and the sample detection concentration range is 10
-15To 10
-3Mole every liter.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201010595059 CN102128938B (en) | 2010-12-20 | 2010-12-20 | Anabolic steroids and polycyclic aromatic hydrocarbon high-efficiency bioluminescence sensor and construction method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 201010595059 CN102128938B (en) | 2010-12-20 | 2010-12-20 | Anabolic steroids and polycyclic aromatic hydrocarbon high-efficiency bioluminescence sensor and construction method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102128938A CN102128938A (en) | 2011-07-20 |
| CN102128938B true CN102128938B (en) | 2013-06-19 |
Family
ID=44267101
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 201010595059 Expired - Fee Related CN102128938B (en) | 2010-12-20 | 2010-12-20 | Anabolic steroids and polycyclic aromatic hydrocarbon high-efficiency bioluminescence sensor and construction method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102128938B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110684792B (en) * | 2019-10-14 | 2022-01-11 | 江南大学 | Glycolic acid sensor capable of being eliminated repeatedly and construction method thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000049150A1 (en) * | 1999-02-18 | 2000-08-24 | National University Of Singapore | Chimeric gene constructs for generation of fluorescent transgenic ornamental fish |
| WO2000071565A2 (en) * | 1999-05-21 | 2000-11-30 | The Regents Of The University Of California | Fluorescent protein indicators |
| CN101539524A (en) * | 2009-03-06 | 2009-09-23 | 于源华 | Fluorescence detection card and fluorescence detection method for organic contaminants in food and environment |
-
2010
- 2010-12-20 CN CN 201010595059 patent/CN102128938B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2000049150A1 (en) * | 1999-02-18 | 2000-08-24 | National University Of Singapore | Chimeric gene constructs for generation of fluorescent transgenic ornamental fish |
| WO2000071565A2 (en) * | 1999-05-21 | 2000-11-30 | The Regents Of The University Of California | Fluorescent protein indicators |
| CN101539524A (en) * | 2009-03-06 | 2009-09-23 | 于源华 | Fluorescence detection card and fluorescence detection method for organic contaminants in food and environment |
Non-Patent Citations (10)
| Title |
|---|
| A model on the regulation of 3α-hydroxysteroid dehydrogenase/carbonyl reductase expression in Comamonas testosteroni;Guangming Xiong等;《Chemico-Biological Interactions》;20010131;第130-132卷;第723-736页 * |
| Cis- and trans-regulatory elements of 3 alpha-hydroxysteroid dehydrogenase/carbonyl reductase as biosensor system for steroid determination in the environment;Guangming Xiong等;《Chemico-BiologicalInteractions》;20091231;第178卷;第215-220页 * |
| Guangming Xiong等.A model on the regulation of 3α-hydroxysteroid dehydrogenase/carbonyl reductase expression in Comamonas testosteroni.《Chemico-Biological Interactions》.2001,第130-132卷 |
| Guangming Xiong等.Cis- and trans-regulatory elements of 3 alpha-hydroxysteroid dehydrogenase/carbonyl reductase as biosensor system for steroid determination in the environment.《Chemico-BiologicalInteractions》.2009,第178卷 |
| 于源华等.环境中甾体激素降解茵的筛选及功能基因的克隆.《长春理工大学学报》.2010,第33卷(第1期), |
| 于源华等.生物荧光方法快速测定环境水体中的甾体类激素.《分析化学》.2010,第38卷(第7期), |
| 液相色谱一串联质谱同时测定动物组织中22种同化激素;陈慧华等;《分析化学》;20090228;第37卷(第2期);第181-186页 * |
| 环境中甾体激素降解茵的筛选及功能基因的克隆;于源华等;《长春理工大学学报》;20100331;第33卷(第1期);第108-111页 * |
| 生物荧光方法快速测定环境水体中的甾体类激素;于源华等;《分析化学》;20100731;第38卷(第7期);第1048-1051页 * |
| 陈慧华等.液相色谱一串联质谱同时测定动物组织中22种同化激素.《分析化学》.2009,第37卷(第2期), |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102128938A (en) | 2011-07-20 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Chien et al. | Enhancing the tropism of bacteria via genetically programmed biosensors | |
| Line et al. | Physiological levels of nitrate support anoxic growth by denitrification of Pseudomonas aeruginosa at growth rates reported in cystic fibrosis lungs and sputum | |
| CN100410362C (en) | Method for culturing chlorella with high-density and high-quality | |
| CN102876579B (en) | Method for culturing and detecting high selenium-enriched spirulina platensis | |
| Wecker et al. | Design of a new biosensor for algal H2 production based on the H2-sensing system of Rhodobacter capsulatus | |
| Rajoka et al. | Kinetics and thermodynamics of ethanol production by a thermotolerant mutant of Saccharomyces cerevisiae in a microprocessor‐controlled bioreactor | |
| UA80809C2 (en) | Process for preparation of protective antigen protein of anthracis with e.coli | |
| CN102191213B (en) | Gene encoding of firefly luciferase, its preparation method and application | |
| Ma et al. | Oxygen transfer rate control in the production of human‐like collagen by recombinant Escherichia coli | |
| CN102128938B (en) | Anabolic steroids and polycyclic aromatic hydrocarbon high-efficiency bioluminescence sensor and construction method thereof | |
| CN106520897A (en) | Method for improving hydrogen production of chlamydomonas | |
| CN102147364B (en) | Efficient microbial fluorescent sensor for detecting steroids hormone and polycyclic aromatic hydrocarbon | |
| CN104877939A (en) | Culture medium and cultural method applied to escherichia coli | |
| Pooley et al. | Continuous culture of photobacterium | |
| CN103122324B (en) | Method for culturing chicken escherichia coli | |
| Nie et al. | Enhanced intracellular glutathione synthesis and excretion capability of Candida utilis by using a low pH‐stress strategy | |
| Zhou et al. | Study on the effect of key genes ME2 and adhE during Luzhou-flavor Baijiu brewing | |
| CN102135497B (en) | High-efficiency chemiluminescence sensor for detecting steroid hormone and polycyclic aromatic hydrocarbon | |
| CN105779354A (en) | Escherichia coli culture medium | |
| CN207958369U (en) | A kind of microbial detection device | |
| CN105349561A (en) | Method of improving fermentation cell density using hemoglobin | |
| CN102818801A (en) | Method for determining ATP | |
| CN101302502B (en) | Method for preparing firefly luciferase | |
| CN108285917A (en) | Detect coliform and the chromogenic culture medium and detection lug of salmonella simultaneously | |
| CN104498391A (en) | Escherichia coli and culture method of culture medium thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C17 | Cessation of patent right | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20130619 Termination date: 20131220 |