CN102321565B - Transgenic salmonella typhimurium TA1535/Pcda-GFP and construction method thereof - Google Patents
Transgenic salmonella typhimurium TA1535/Pcda-GFP and construction method thereof Download PDFInfo
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Abstract
The present invention discloses transgenic salmonella typhimurium TA1535/Pcda-GFP and a construction method thereof. The preservation number of the transgenic salmonella typhimurium TA1535/Pcda-GFP is CCTCC NO: M 2011269. According to the construction method, recombinant plasmid with a strong promoter Pcda and green fluorescent protein gene EGFP is transformed into salmonella typhimurium TA1535 to obtain the transgenic salmonella typhimurium TA1535/Pcda-GFP. Compared to the wild type salmonella typhimurium TA1535, the engineered strain provided by the present invention provides enhanced fluorescence intensity, thus, when the engineered strain is adopted for a toxicity test, the sensitivity of the toxicity test method is improved, the toxic material can be detected conveniently and rapidly. Therefore, the transgenic salmonella typhimurium TA1535/Pcda-GFP provided by the present invention provides convenience for realization of convenient, rapid and sensitive detection of the toxic material, and can be applicable for popularization and application in detection of the genetic material in the environment.
Description
Technical field:
The invention belongs to biological technical field, be specifically related to a kind of transgenic salmonella typhimurium TA 1535 TA1535/Pcda-GFP engineering strain and construction process thereof.
Background technology:
Traditional toxicity test method such as pilot fish, algae experiment, zooplankton test etc., or expense is relatively costly, or the complex operation test is consuming time longer, is unfavorable for rapid detection or on-line monitoring.And the photogenic bacterium rule can well overcome these defectives, and it has fast, sensitivity and simple operation and other advantages.The application photogenic bacterium carries out toxotest can the following aspects: one is to utilize natural photogenic bacterium to carry out toxotest, bacterial luminescence intensity becomes negative correlativing relation (being called light-off) with toxicant concentration, with luminous inhibiting rate reflection comprehensive toxicity size, this method is widely accepted; Use recombinant DNA technology and make up restructuring photogenic bacterium (Recombinant Luminescent Bacteria), be divided into two kinds of composing type and induction types, wherein induction type must just be expressed by luciferase when specific inductor exists, in poisonous substance sublethal concentrations scope, its dose-effect relationship is positive correlation (showing as light-on); Also have is exactly to utilize to extract to such an extent that enzyme system is carried out toxicity test from luminous organism.
Traditional photogenic bacterium method can only provide the comprehensive toxicity of sample, and composition gets over complicated municipal effluent and trade effluent requires not only will reflect in the surroundings monitoring the total toxicity size of water body, and the information of dissimilar toxicants more will be provided.So the restructuring photogenic bacterium begins to enter people's the visual field.
The lux operon is as coding and the luminous gene of regulation and control bacterium living beings, and the direct result of its expression is very directly perceived and be convenient to the instrument detection, therefore is widely used as reporter gene with the research gene expression and regulation in molecular biology research.Based on these characteristics of lux reporter gene, just begin to have the scholar to propose to make up the method that the restructuring photogenic bacterium comes specific toxicant in the testing environment the nineties in last century.Photogenic bacterium is compared with natural photogenic bacterium in restructuring, it is advantageous that different regulatory gene and promotor given RLB different recognition capabilities, can be according to all kinds of toxicants of the specific identification of toxic effect mechanism, so can be in order to detecting various cell injury, or certain class special compound such as environment incretion interferent (EDCs), persistence organic pollutant (POPs) etc. in the testing environment.The nearly more than ten years, along with the concern of people to RLB, more and more various RLB with difference in functionality are created out.
The SOS/Umu test macro just is being based on the dna damage thing and is inducing the SOS reaction and express the ability of umuC gene, in Salmonella typhimurtum Salmonella typhimuriumTA1535, import and carry umuC " the chimeric plasmid pSK1002 of LacZ; this plasmid carries the umu operon; umuD gene and umuC " promotor and tsiklomitsin and the paraxin drug resistant gene of LacZ fusion gene, allow by the medicament selection transformant, the bacterium of new structure is called S.typhimurium TA1535-pSK1002, and " when bacterium is subjected to the paramorphogen effect to cause the SOS reaction; activate the umuC-LacZ fusion gene; give expression to the fusion rotein of betagalactosidase activity; can determine that by the activity that detects this enzyme tested material causes the degree of dna damage; if add the S9 mixed solution, whether metabolism activation generates the product that damages DNA then can to detect chemicals.
Summary of the invention:
The purpose of this invention is to provide a kind of more convenient, sensitive, monitor transgenic salmonella typhimurium TA 1535 (Salmonella typhimurium) TA1535/Pcda-GFP engineering strain and the construction process thereof of toxicant fast.
Transgenic salmonella typhimurium TA 1535 of the present invention (Salmonella typhimurium) TA1535/Pcda-GFP was preserved in Chinese Typical Representative culture collection center (CCTCC) on 07 26th, 2011, the address: Wuhan, China city Wuhan University, its deposit number is CCTCC NO:M2011269.
Salmonella bacillus typhi murium of the present invention (Salmonella typhimurium) TA1535/Pcda-GFP makes up by the following method, may further comprise the steps:
Strong promoter Pcda is connected with green fluorescence protein gene EGFP, again with expression vector pET20b(+) be connected and be built into recombinant plasmid PSK-EGFP, be transformed into again among Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535, thereby obtain genetically modified Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535/Pcda-GFP, the nucleotide sequence of described strong promoter Pcda is shown in 98-219 of SEQ ID NO.1, and the nucleotide sequence of described green fluorescence protein gene EGFP is shown in 220-936 of SEQ ID NO.1.
Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535 that the present invention uses belongs to bacterial strain of the prior art, can buy from German DSMZ DSMZ.
The present invention will be transferred to the recombinant plasmid of strong promoter Pcda and green fluorescence protein gene EGFP among the Salmonella bacillus typhi murium TA1535, obtain therefrom genetically modified Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535.This project bacterial strain is than the Salmonella bacillus typhi murium TA1535 of wild-type, and its fluorescence intensity strengthens greatly, thereby when it was used for toxotest, the sensitivity that can improve the toxotest method can detect toxicant more convenient, faster.Therefore of the present inventionly be embodied as more convenient, quick, sensitive detection toxicant and provide convenience, can be in environment be applicable in the detection of genetic material.
Salmonella bacillus typhi murium of the present invention (Salmonella typhimurium) TA1535/Pcda-GFP was preserved in Chinese Typical Representative culture collection center (CCTCC) on 07 26th, 2011, the address: Wuhan, China city Wuhan University, its deposit number is CCTCC NO:M2011269.
Description of drawings:
Fig. 1 is strong promoter Pcda pcr amplification qualification result, M:DL2000DNAMarker, Pcda:Pcda amplified production, 2, negative control;
Fig. 2 is green fluorescence protein gene EGFP pcr amplification qualification result, M:DL5000DNAMarker, EGFP, EGFP amplified production, 2, negative control;
Fig. 3 is the pcr amplification qualification result after Pcda-EGFP connects, M:DL2000DNAMarker, 1,2, negative control, 3,4, Pcda-EGFP connects product;
Fig. 4 is hickie pure culture figure;
Fig. 5 is hickie pure culture burst of ultraviolel figure, and excitation wavelength is 365nm;
Fig. 6 is the salmonella typhi TA1535(1 and 2 of wild-type) and transgenosis salmonella typhi TA1535/Pcda-GFP(sun 1 and sun 2) the burst of ultraviolel figure of engineering strain, excitation wavelength is 365nm.
Embodiment:
Following examples are to further specify of the present invention, rather than limitation of the present invention.
Embodiment 1:
One, the amplification of strong promoter Pcda:
Give birth to the plasmid that contains the Pcda promotor (Description of Goods is as Pcda) that worker's biotechnology Services Co., Ltd provides take Shanghai as template, take C1 and C2 as the upstream and downstream primer, the PCR reaction system is (Pcda promoter plasmid 100ng, 1 μ L respectively; C150 μ M, 1 μ L; C250 μ M, 1 μ L; DNTP (10mM) 1 μ L; Taq enzyme 5-10U, 1 μ L; Low salt buffer, 5 μ L; Deionized water, 45 μ L), carry out PCR reaction (95 ℃ of 4min, 95 ℃ of 45s, 52 ℃ of 30s, 72 ℃ of 30s, 29cycle, 72 ℃ of 10min, 22 ℃ of forever), the PCR product shows that with 1% agarose electrophoresis the clip size that increases is (Fig. 1) about 120bp, conform to bibliographical information, the result meets expection.
C1：N:5′-AGCGAATTCGGGTTGACAGGATTTACG-3′
C2：C:5′-TTCTCCTTTACTCATGCAAACACCT-3′
Two, the amplification of green fluorescence protein gene EGFP:
With the plasmid that contains green fluorescence protein gene EGFP (Shanghai Ya Ji bio tech ltd, article No.: YJ-2194P) be template, take P1 and P2 as the upstream and downstream primer, the PCR reaction system is (EGFP plasmid, 1 μ L respectively; P150 μ M, 1 μ L; P250 μ M, 1 μ L; DNTP (10mM) 1 μ L; Taq enzyme 5-10U, 1 μ L; Low salt buffer, 5 μ L; Deionized water, 45 μ L), carry out PCR reaction (95 ℃ of 4min, 95 ℃ of 45s, 56 ℃ of 45s, 72 ℃ of 1min, 34cycle, 72 ℃ of 10min, 22 ℃ of forever), the PCR product shows that with 1% agarose electrophoresis the clip size that increases is (Fig. 2) about 750bp, and the result meets expection.
P1N:5′-ATGAGTAAAGGAGAAGAAC-3′
P2C:5′-CTA
CTTGATATGTTTATTTTCGAACGTGA-3′
Three, the Pcda-EGFP fragment connects:
Respectively top pcr amplification product (strong promoter Pcda fragment and green fluorescence protein gene EGFP fragment) is carried out glue and reclaim purifying, by OD260/OD280, determine DNA concentration, concentration is respectively 168ng/ μ L and 243ng/ μ L.Process purifying Pcda and EGFP fragment are carried out without primer PCR, and the PCR reaction system is (strong promoter Pcda fragment 100ng, 1 μ L; Green fluorescence protein gene EGFP fragment 100ng, 1 μ L, dNTP (10mM) 1 μ L; Taq enzyme 5-10U, 1 μ L; Low salt buffer, 5 μ L; Deionized water, 45 μ L), the PCR program is: 94 ℃ of 40s, 48 ℃ of 40s, 72 ℃ of 2min, 15cycle.Then with C1 and P2 as the upstream and downstream primer, the PCR reaction system is (strong promoter Pcda fragment 100ng, 1 μ L; Green fluorescence protein gene EGFP fragment 100ng, 1 μ L; C150 μ M, 1 μ L; P250 μ M, 1 μ L; DNTP (10mM) 1 μ L; Taq enzyme 5-10U, 1 μ L; Low salt buffer, 5 μ L; Deionized water, 45 μ L), carry out pcr amplification (94 ℃ of 4min, 94 ℃ of 40s, 54 ℃ of 1min, 72 ℃ of 1min, 30cycle, 22 ℃ of forever), the PCR product shows that through 1% agarose electrophoresis the clip size that increases is (Fig. 3) about 900bp.
Reclaim the Pcda-EGFP fragment by cutting glue.Behind the purifying, our purpose fragment is connected to TA cloning vector PMD18, checks order.Sequencing result is shown in SEQ ID NO.1, and through comparison, its 98-219 is strong promoter Pcda, and 220-936 is green fluorescence protein gene EGFP.Sequencing result is consistent with the experimental design fragment.Sequencing result shows that promotor Pcda and green fluorescence protein gene EGFP correctly insert, and successful connection obtains the Pcda-EGFP junction fragment thus.
Four, Pcda-EGFP fragment and expression vector pET20b(+) (a lucky bio tech ltd is wished in Shanghai, article No.: CPC15) connect:
1, enzyme is cut
With 1 μ g Pcda-EGFP fragment, EcoR I0.5 μ L, HindIII0.5 μ L, 10 * M Buffer2 μ L adds sterilized water to 20 μ L, and 37 ℃ of (18h) enzymes that spend the night are cut.
With 1 μ g pET20b(+), EcoR I0.5 μ L, HindIII0.5 μ L, 10 * M Buffer2 μ L adds sterilized water to 20 μ L, and 37 ℃ of (18h) enzymes that spend the night are cut.
2, connect
T4 enzyme 1 μ L, 10 * T4DNA Ligase Buffer2.5 μ L, expression vector pET20b(+) enzyme cut product 1 μ L, it is 3 μ L that the enzyme of Pcda-EGFP fragment is cut product.Add sterilized water to 25 μ L, 16 ℃ spend the night make Pcda-EGFP fragment and expression vector pET20b(+) be connected, therefrom with Pcda-EGFP fragment inserting expressioning carrier pET20b(+) in, make up and to obtain recombinant plasmid PSK-EGFP.
Five, recombinant plasmid PSK-EGFP is changed among the E.coli TOP10 preserve
Get above-mentioned 90 μ l recombinant plasmid PSK-EGFP and add electric shock cup bottom, add again E.coli TOP10 bacterial strain, touch liquid and be positioned at electric shock cup bottom to guarantee bacterium and DNA suspension, dry electric shock tank outer water of condensation and fog, the cup that will shock by electricity is put the electric shock instrument conversion of shocking by electricity, 2500v, 5ms into.After electric shock finished, the cache sample pool added 1mL SOC nutrient solution under the room temperature as far as possible, and 37 ℃, 200rpm/min cultivates 1h.Get respectively 20 μ l, 50 μ l, 100 μ l coat on the ammonia benzyl flat board that contains 50mg/L.Be inverted for 37 ℃ and cultivate, place 10-16h.Choose hickie, as primer, use verify (Fig. 3) of PCR method with C1 and P2.Obtain therefrom the E.coli TOP10 recombinant bacterial strain with recombinant plasmid PSK-EGFP.
Six, the structure of Salmonella bacillus typhi murium (salmonella typhimurium) TA1535/Pcda-GFP
Extraction is with the plasmid of the E.coli TOP10 recombinant bacterial strain of recombinant plasmid PSK-EGFP, after extracting is finished, use the RNA enzyme, processed one hour for 37 ℃, with the LB incubated overnight without the Salmonella bacillus typhi murium TA1535 bacterial strain of ammonia benzyl resistance (since in the Salmonella bacillus typhi murium TA1535 bacterial strain with the Psk1002 plasmid, will inevitably exert an influence to ammonia benzyl resistance screening, therefore can carry out to the Salmonella bacillus typhi murium TA1535 bacterial strain of wild-type continuous passage cultivates, the Psk1002 plasmid is lost, therefrom screen the Salmonella bacillus typhi murium TA1535 bacterial strain without ammonia benzyl resistance), according to the preceding method conversion of shocking by electricity, after the conversion, choose bacterium Ultraluminescence checking.
Shown in Fig. 4,5,6, Fig. 5 and Fig. 6 are burst of ultraviolel figure, adopt the 365nm wavelength, the Salmonella bacillus typhi murium TA1535 that contains recombinant plasmid PSK-EGFP that transforms successfully excites down ultraviolet, thalline sends green fluorescence, 28 bacterium colonies in Fig. 5 have all sent green fluorescence, this shows, these bacterial strains have all transformed successfully, as shown in Figure 6, transform successful positive colony and send green fluorescence, and the Salmonella bacillus typhi murium TA1535 without ammonia benzyl resistance of wild-type does not send fluorescence.Can be inferred by above-mentioned figure, the TA1535 recombinant bacterial strain (positive colony) that contains recombinant plasmid PSK-EGFP successfully constructs, with this bacterium called after Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535/Pcda-GFP, this bacterium was preserved in Chinese Typical Representative culture collection center (CCTCC) on 07 26th, 2011, the address: Wuhan, China city Wuhan University, its deposit number is CCTCC NO:M2011269.
Function ratio
Picking Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535/Pcda-GFP(CCTCC NO:M2011269) and single bacterium colony of the Salmonella bacillus typhi murium TA1535 of wild-type method:, with LB+Amp(50 μ g/mL) liquid nutrient medium carries out 30 ° of C, and 200rpm spends the night and shakes bacterium.With the 1:50(volume ratio) be transferred to contain 50mL(LB+50 μ g/mLAmp) Erlenmeyer flask in, 30 ° of C, 200rpm shakes bacterium.Control is shaken the bacterium time, gets different time sections bacterium liquid and carries out exposure experiment, measures OD
600Be respectively 0.8, the reaction times is 6h, gets 3mL as sample, carries out spectrophotofluorometer and carries out fluorescent strength determining.Positive material adopts ametycin (MMC), and concentration is 1 μ g/mL, and negative control compares the relative intensity of fluorescence under the same terms with dimethyl sulfoxide (DMSO) (DMSO).
The results are shown in Table 1:
Table 1: two bacterial classifications fluorescence intensity under the same conditions
As can be seen from Table 1, the fluorescence intensity of Salmonella bacillus typhi murium of the present invention (Salmonella typhimurium) TA1535/Pcda-GFP under the same conditions, its fluorescence intensity (26.3) is far longer than the Salmonella bacillus typhi murium TA1535(11.8 of wild-type), when therefore using it for toxotest, the sensitivity of toxotest method can be improved, toxicant can be detected more convenient, faster.Therefore of the present inventionly be embodied as more convenient, quick, sensitive detection toxicant and provide convenience, can be in environment be applicable in the detection of genetic material.
Claims (1)
1. the construction process of a Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535/Pcda-GFP, it is characterized in that, may further comprise the steps: strong promoter Pcda is connected with green fluorescence protein gene EGFP, again with expression vector pET20b(+) be connected and be built into recombinant plasmid PSK-EGFP, be transformed into again among Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535, thereby obtain genetically modified Salmonella bacillus typhi murium (Salmonella typhimurium) TA1535/Pcda-GFP, the nucleotide sequence of described strong promoter Pcda is shown in 98-219 of SEQ ID NO.1, the nucleotide sequence of described green fluorescence protein gene EGFP is shown in 220-936 of SEQ ID NO.1, and the deposit number of this bacterium is: CCTCC NO:M2011269.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1448504A (en) * | 2003-05-07 | 2003-10-15 | 华东师范大学 | Construction and application of engineering bacterium LZ-17 |
| CN101255456A (en) * | 2008-04-01 | 2008-09-03 | 中国烟草总公司郑州烟草研究院 | Toxicology neasuring evaluation method of cigarette smoke mutagenicity |
| CN101851659A (en) * | 2010-04-23 | 2010-10-06 | 云南烟草科学研究院 | Ames experiment micro fluctuation method of mutagenicity detection for water bodies |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1448504A (en) * | 2003-05-07 | 2003-10-15 | 华东师范大学 | Construction and application of engineering bacterium LZ-17 |
| CN101255456A (en) * | 2008-04-01 | 2008-09-03 | 中国烟草总公司郑州烟草研究院 | Toxicology neasuring evaluation method of cigarette smoke mutagenicity |
| CN101851659A (en) * | 2010-04-23 | 2010-10-06 | 云南烟草科学研究院 | Ames experiment micro fluctuation method of mutagenicity detection for water bodies |
Non-Patent Citations (4)
| Title |
|---|
| Construction of a ColD cda Promoter-Based SOS-Green Fluorescent Protein Whole-Cell Biosensor with Higher Sensitivity toward Genotoxic Compounds than Constructs Based on recA,umuDC, or sulA Promoters;Norman A., et al.;《Applied And Environment Microbiology》;20050531;第71卷(第5期);2338－2346 * |
| NormanA. et al..Construction of a ColD cda Promoter-Based SOS-Green Fluorescent Protein Whole-Cell Biosensor with Higher Sensitivity toward Genotoxic Compounds than Constructs Based on recA |
| 刘薇，等.用SOS/umu测试评价光电催化降解五氯酚过程的遗传毒性特征.《生态毒理学报》.2006,第1卷(第2期),155－159. * |
| 张家强,等.水源水中的遗传毒性检测技术研究进展.《环境科学与技术》.2010,第33卷(第6E期),188-192. * |
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