CN103710335B - A kind of method utilizing atmospheric pressure at room plasma body to prepare microorganism competent - Google Patents
A kind of method utilizing atmospheric pressure at room plasma body to prepare microorganism competent Download PDFInfo
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- CN103710335B CN103710335B CN201310700136.9A CN201310700136A CN103710335B CN 103710335 B CN103710335 B CN 103710335B CN 201310700136 A CN201310700136 A CN 201310700136A CN 103710335 B CN103710335 B CN 103710335B
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Abstract
The invention discloses a kind of method utilizing atmospheric pressure at room plasma body to prepare microorganism competent, belong to the application of plasma technique at biological technical field.Ultimate principle of the present invention utilizes energetic ion in plasma body and charged particle, interact with the protein in microorganism cells and genetic material, finally cause the change of cellularity, make cell be in the suitableeest picked-up and hold the physiological status of foreign DNA.The present invention adopts atmospheric pressure at room plasma body in conjunction with fluid circulation system, take helium as plasma discharge gas, acts on microorganism cells, efficiently, easily can prepare competent cell.The advantages such as the present invention has efficiently, quick, safe, convenient, can be used as a kind of Novel experimental technology and are widely used in microorganism hereditary, molecular genetic, genetically engineered field.
Description
Technical field
The invention belongs to the application of plasma technique in field of biology, be specifically related to a kind of method utilizing atmospheric pressure at room plasma body to prepare microorganism competent.
Background technology
The process making the DNA molecular with genetic information enter host cell is called " conversion "; Through physics or chemical mode process and there is the host cell that acceptant additional DNA segments enters, be called " competent cell ".The state of competent cell directly affects the efficiency of conversion, and in order to make host cell carry out copying or expressing of specific DNA or protein, it is crucial for obtaining the competent cell being easy to transform.
The method preparing competent cell mainly comprises physical method and the large class of chemical process two.Wherein comparatively conventional physical method comprises electricity irritation, ultrasonic wave, osmotic pressure etc.; Chemical process comprises CaCl
2, RbCl (KCl), Inoue method etc.But often there is the problems such as transformation efficiency is lower, treating processes is loaded down with trivial details, environmental pollution in chemical process, and the general energy consumption of physical method is higher, cells survival rate is low.Therefore, study high, the simple to operate practicality of a kind of transformation efficiency and method with low cost for the preparation of competent cell, particularly necessary to molecular biological research.
Plasma body is the 4th state of material, and by molecular systems of grain such as a large amount of molecules, atom, ion and electronics, chemically reactive is high and be macroscopically electric neutrality.The technology that this mild condition, energy density are high is applied to the preparation of competent cell gradually, as invention " a kind of method utilizing atmosphere cold plasma to improve permeability of cell membrane " (notification number: CN102321606A), namely atmospheric dielectric barrier discharge (Atmospheric-PressureDielectricBarrierDischarge is utilized, APDBD) Cement Composite Treated by Plasma microorganism cells suspension, reaches the object improving permeability of cell membrane.But DBD electric discharge is even not, makes its application be restricted.
Atmospheric pressure at room plasma body (AtmosphericandRoomTemperaturePlasma, ARTP) owing to eliminating dielectric overlay, electric discharge is more even compared with APDBD, and generator architecture is simple, cost is low, produce the temperature near room temperature of plasma jet, and ozone concn and ultraviolet radiation intensity are very low.ARTP is rich in helium atom, Sauerstoffatom, nitrogen-atoms, the OH free radical homenergic ion of excited state, can interact with the protein in microorganism cells and genetic material, finally cause the change of cellularity, make cell be in the suitableeest picked-up and hold the physiological status of foreign DNA.But, there is not the report about utilizing atmospheric pressure at room plasma body to prepare microorganism competent in prior art.
Summary of the invention
The object of the present invention is to provide a kind of method utilizing atmospheric pressure at room plasma body to prepare microorganism competent.The method has simple, convenient, efficient, economic, practical feature, is specially adapted to the preparation of microorganism competent.
Technical scheme of the present invention is as follows:
Utilize atmospheric pressure at room plasma body to prepare a method for microorganism competent, it is characterized in that, during preparation, make microorganism cells suspension through plasma irradiating, collect, obtain competent cell.
Described plasma body is atmospheric pressure at room plasma body, the uniform glow discharge plasma body adopting bare metal electrode to produce under normal atmosphere, room temperature condition by ARTP selection by mutation system.
Described ARTP selection by mutation system, with helium, argon gas, nitrogen, oxygen, air or its mixed gas for source of the gas.
The parameter of described ARTP selection by mutation system is: described source of the gas is helium, argon gas or its mixed gas are source of the gas, and gas flow is set as 3 ~ 30SLM(StandardLiterperMinute, public liter/min), power is 50 ~ 150W.In this parameter area, plasma discharge is evenly effective.
Described gas flow is set as that 5 ~ 15SLM, power are 80 ~ 120W, and in this parameter area, plasma discharge uniformity coefficient is best, and effect is best.
Described fluid circulation system adjusts flow velocity by peristaltic pump, and each circular treatment time is 0 ~ 20s, and this concentration range can meet the requirement preparing competent cell substantially.
Each circular treatment time is 5 ~ 15s, both can obtain a large amount of competent cells within the scope of this, cell membrane can not cause damage again; Described microorganism cells suspension is bacterial cell suspension, actinomycetes cells suspension, fungal cell's suspension, alga cells suspension or algae protoplasm somatocyte suspension.
Aforesaid method comprises the steps:
1) select the single colony inoculation of microorganism of new activation in liquid nutrient medium from the solid plate of microorganism growth, concussion overnight incubation, obtains bacterium liquid;
2) be inoculated in the new liquid nutrient medium changed by the bacterium liquid that step 1) obtains, concussion is cultured to bacterium liquid OD
600be 0.3 ~ 1.0, this concentration range can meet the requirement accepting plasma irradiating equably substantially.
3) by step 2) concussion gained bacterium liquid adds in sterile centrifugation tube, centrifugal, and abandon supernatant, retain precipitation thalline;
4) with aseptic 0.1mol/LCaCl
2the thalline of the resuspended step 3) centrifugation of the aqueous solution;
5) with described plasma treatment step 4) thalline that obtains;
6) collect the thalline of step 5) after Cement Composite Treated by Plasma, be competent cell.
Step 2) in, described bacterium liquid OD
600be 0.5 ~ 0.7, this scope inner cell moderate concentration, more uniformly can accept plasma irradiating;
The inoculum size of described bacterium liquid is 5% ~ 15%, to provide more suitable growth concentration.
In step 3), described is centrifugal, rotating speed 4000 ~ 10000r/min, centrifugation time 0.5 ~ 2.0min, with abundant sample separation, and unlikely cell wound again.
The invention is characterized in and carry out according to the following steps successively:
(1) bacteria suspension is prepared: select the single colony inoculation of microorganism of new activation in liquid seed culture medium from the solid plate of applicable microorganism growth, under the optimum growing condition of microorganism, shake overnight incubation; By the bacteria suspension of acquisition by 10% inoculum size transfer in fresh liquid seed culture medium, under the optimum growth temperature of this microorganism concussion be cultured to OD
600be between 0.5 ~ 0.7; By centrifugal for the bacteria suspension loading sterile centrifugation tube obtained, supernatant discarded, use 0.1mol/LCaCl
2the aqueous solution is resuspended.
(2) optimum configurations: using helium as plasma discharge gas, helium gas flow is set as 5 ~ 15SLM, power 80 ~ 120W.
(3) described pending bacterium liquid is added in stoste storage bottle, bacterium liquid sends into overflow device bottom liquid inlet mouth through pipeline by peristaltic pump, and bacterium liquid is spilled in accumulator tank by overflow port to surrounding, and treatment solution flows into treatment liquid storage bottle by recovery tube.Overflow device is placed in plasma generator fluerics, at room temperature environment process 5s ~ 30min, and each circular treatment time 5 ~ 15s.
(4) bacterium liquid in storage bottle is moved to aseptic triangular flask, add sterile glycerol, make the final content of glycerine reach volume percent 15% ~ 20%, the frozen competent cell of packing.
Plasma body described in the present invention is used for the Efficient Conversion of multiple plasmid and knocking out of chromogene.
The inventive method has efficiently, fast, feature easily.
Accompanying drawing explanation
Fig. 1 utilizes plasma body to prepare the method flow diagram of microorganism competent
Embodiment
Following embodiment is provided to be to understand the present invention further better; be not limited to described preferred forms; content of the present invention and protection domain are not construed as limiting; anyone is under enlightenment of the present invention or any and the present invention the present invention being carried out combining with the feature of other prior aries and draw is identical or akin product, all drops within protection scope of the present invention.
Unreceipted specific experiment step or condition person in embodiment, can carry out according to the operation of the normal experiment step described by the document in this area or condition.Agents useful for same or the unreceipted production firm person of instrument, being can by the conventional reagent product of commercial acquisition.
ARTP selection by mutation system is that Beijing Siqingyuan Bioscience Co., Ltd. prepares according to the patent of invention of ZL201110339695.2.
embodiment 1: the preparation of intestinal bacteria Top10 bacterial strain competent cell and transformation efficiency measure
1, material and reagent
Material: recipient bacterium E.coliTOP10; Plasmid: pUC
Reagent: LB liquid nutrient medium: peptone 10g, yeast extract paste 5g, NaCl10g, distilled water constant volume 1L, pH7.0,121 DEG C of sterilizing 20min.
LB solid medium: agar 10g, peptone 10g, yeast extract paste 5g, NaCl10g, distilled water constant volume 1L, pH7.0,121 DEG C of sterilizing 20min.
2, laboratory apparatus
ARTP(atmospheric pressure at room plasma body) selection by mutation system
3, experimental procedure:
1) treatment solution preparation: picking list colony inoculation is in 5mLLB liquid nutrient medium on the solid plate of growth Top10 bacterial strain, cultivates 12 ~ 16 hours for 37 DEG C; Get 2mL culture switching kind of (inoculum size is 5%) to continue to grow to bacterium liquid OD in the fresh LB liquid nutrient medium of 200mL
600be 0.5; Bacterium liquid is sub-packed in the centrifugal 1min of 4000r/min in the freezing centrifuge tube of 50mL, reclaims cell; Abandon most supernatant, by bacterial precipitation Eddy diffusion in 20mL0.1mol/LCaCl
2the aqueous solution.
2) set handling parameter: with high-purity helium for source of the gas, gas flow 10SLM, power 120W, process distance 2mm, flow velocity 5rpm, carry out a circulation, cycling time is 5s.
3) acquisition of competent cell: the bacteria suspension after collection and treatment is competent cell.
4) mensuration of transformation efficiency: get 1 μ LpUC plasmid DNA and add in 200 μ L competent cells; Gently after mixing, ice bath 30min; 42 DEG C, heat-shocked 90s, is then placed in rapidly ice bath cooling 3min; Add 1mLLB liquid nutrient medium, 37 DEG C of shaking culture 2h, make Host Strains recover and express resistance protein.By centrifugal for bacterium liquid, abandon supernatant, it is resuspended to add 200 μ L fresh bacterium liquid, gets 100 μ L and coats corresponding LB solid medium.Be inverted cultivation 12 ~ 16h for 37 DEG C, after bacterium colony grows, colony count, calculates transformation efficiency.By this experiment in triplicate, do 3 conversions, calculating mean value, determines transformation efficiency at every turn.
Table 1. is through the transformation efficiency of different time atmospheric pressure at room Cement Composite Treated by Plasma intestinal bacteria Top10
| Time (s) | 0 | 15 | 30 | 45 | 60 | 75 |
| Transformation efficiency (%) | 0 | 51 | 34 | 5 | 2 | 1 |
As shown in table 1, within the scope of 15 ~ 75s, thalline transformation efficiency reduces gradually with the increase in treatment time, and the treatment time needed for transformation efficiency reaching 51% is only 15s, illustrates that the method fast, efficiently.
embodiment 2: the preparation of bacillus coli DH 5 alpha competent cell and transformation efficiency measure
1, material and reagent
Material: recipient bacterium E.coliDH5 α; Plasmid pUC
Reagent: LB liquid nutrient medium: peptone 10g, yeast extract paste 5g, NaCl10g, distilled water constant volume 1L, pH7.0,121 DEG C of sterilizing 20min.
LB solid medium: agar 10g, peptone 10g, yeast extract paste 5g, NaCl10g, distilled water constant volume 1L, pH7.0,121 DEG C of sterilizing 20min.
2, laboratory apparatus
ARTP(atmospheric pressure at room plasma body) selection by mutation system
3, experimental procedure
1) treatment solution preparation: picking list colony inoculation is in 5mLLB liquid nutrient medium on the solid plate of growth DH5 α bacterial strain, cultivates 12 ~ 16 hours for 37 DEG C; Get 2mL culture switching kind of (inoculum size is 10%) to continue to grow to bacterium liquid OD in the fresh LB liquid nutrient medium of 200mL
600be 0.6; Bacterium liquid is sub-packed in the centrifugal 1min of 6000r/min in the freezing centrifuge tube of 50mL, reclaims cell; Abandon most supernatant, by bacterial precipitation Eddy diffusion in 20mL0.1mol/LCaCl
2the aqueous solution.
2) set handling parameter: with high-purity helium for source of the gas, gas flow 5SLM, power 100W, process distance 3mm, flow velocity 5rpm, carry out a circulation, cycling time is 10s.
3) acquisition of competent cell: the bacteria suspension after collection and treatment is competent cell.
4) mensuration of transformation efficiency: get 1 μ LpUC plasmid DNA and add in 200 μ L competent cells; Gently after mixing, ice bath 30min; 42 DEG C, heat-shocked 90s, is then placed in rapidly ice bath cooling 3min; Add 1mLLB liquid nutrient medium, 37 DEG C of shaking culture 2h, make Host Strains recover and express resistance protein.By centrifugal for bacterium liquid, abandon supernatant, it is resuspended to add 200 μ L fresh bacterium liquid, gets 100 μ L and coats corresponding LB solid medium.Be inverted cultivation 12 ~ 16h for 37 DEG C, after bacterium colony grows, colony count, calculates transformation efficiency.By this experiment in triplicate, do 3 conversions, calculating mean value, determines transformation efficiency at every turn.
Table 2. is through the transformation efficiency of different time atmospheric pressure at room Cement Composite Treated by Plasma bacillus coli DH 5 alpha
| Time (s) | 0 | 15 | 30 | 45 | 60 | 75 |
| Transformation efficiency (%) | 0 | 64 | 28 | 16 | 8 | 1 |
As shown in Table 2, within the scope of 15 ~ 75s, along with the treatment time increases, thalline transformation efficiency reduces gradually, and when being 15s when treated, transformation efficiency can reach 64%.
embodiment 3: the preparation of intestinal bacteria Top10 bacterial strain competent cell and transformation efficiency measure
1, material and reagent
Material: recipient bacterium E.coliTOP10; Plasmid: pUC
Reagent: LB liquid nutrient medium: peptone 10g, yeast extract paste 5g, NaCl10g, distilled water constant volume 1L, pH7.0,121 DEG C of sterilizing 20min.
LB solid medium: agar 10g, peptone 10g, yeast extract paste 5g, NaCl10g, distilled water constant volume 1L, pH7.0,121 DEG C of sterilizing 20min.
2, laboratory apparatus
ARTP(atmospheric pressure at room plasma body) selection by mutation system
4, experimental procedure:
1) treatment solution preparation: picking list colony inoculation is in 5mLLB liquid nutrient medium on the solid plate of growth Top10 bacterial strain, cultivates 12 ~ 16 hours for 37 DEG C; Get 2mL culture switching kind of (inoculum size is 15%) to continue to grow to bacterium liquid OD in the fresh LB liquid nutrient medium of 200mL
600be 0.5 ~ 0.7; Bacterium liquid is sub-packed in the centrifugal 1min of 8000r/min in the freezing centrifuge tube of 50mL, reclaims cell; Abandon most supernatant, by bacterial precipitation Eddy diffusion in 20mL0.1mol/LCaCl
2the aqueous solution.
2) set handling parameter: with high-purity helium for source of the gas, gas flow 15SLM, power 80W, process distance 4mm, flow velocity 5rpm, carry out a circulation, cycling time is 15s.
3) acquisition of competent cell: the bacteria suspension after collection and treatment is competent cell.
4) mensuration of transformation efficiency: get 1 μ LpUC plasmid DNA and add in 200 μ L competent cells; Gently after mixing, ice bath 30min; 42 DEG C, heat-shocked 90s, is then placed in rapidly ice bath cooling 3min; Add 1mLLB liquid nutrient medium, 37 DEG C of shaking culture 2h, make Host Strains recover and express resistance protein.By centrifugal for bacterium liquid, abandon supernatant, it is resuspended to add 200 μ L fresh bacterium liquid, gets 100 μ L and coats corresponding LB solid medium.Be inverted cultivation 12 ~ 16h for 37 DEG C, after bacterium colony grows, colony count, calculates transformation efficiency.By this experiment in triplicate, do 3 conversions, calculating mean value, determines transformation efficiency at every turn.
Table 3. is through the transformation efficiency of different time atmospheric pressure at room Cement Composite Treated by Plasma intestinal bacteria Top10
| Time (s) | 0 | 15 | 30 | 45 | 60 | 75 |
| Transformation efficiency (%) | 0 | 62 | 30 | 10 | 7 | 1 |
As shown in Table 3, within the scope of 15 ~ 75s, along with the treatment time increases, thalline transformation efficiency reduces gradually, and when being 15s when treated, transformation efficiency can reach 62%.This shows, utilizes atmospheric pressure at room plasma body can prepare competent cell fast and efficiently.
Last it is noted that above embodiment is only in order to illustrate technical scheme of the present invention, be not intended to limit; Although with reference to previous embodiment to invention has been detailed description, those of ordinary skill in the art is to be understood that: it still can be modified to the technical scheme described in foregoing embodiments, or carries out equivalent replacement to wherein portion of techniques feature; And these amendments or replacement, do not make the essence of appropriate technical solution depart from the spirit and scope of various embodiments of the present invention technical scheme.
Claims (8)
1. utilize atmospheric pressure at room plasma body to prepare a method for microorganism competent, it is characterized in that, during preparation, make microorganism cells suspension through plasma irradiating, collect, obtain competent cell; Described plasma body is atmospheric pressure at room plasma body, the uniform glow discharge plasma body adopting bare metal electrode to produce under normal atmosphere, room temperature condition by ARTP selection by mutation system.
2. method according to claim 1, is characterized in that, described ARTP selection by mutation system, with helium, argon gas, nitrogen, oxygen, air or its mixed gas for source of the gas.
3. method according to claim 1, is characterized in that, the parameter of described ARTP selection by mutation system is: source of the gas is helium, argon gas or its mixed gas, and gas flow is set as 3 ~ 30SLM, and power is 50 ~ 150W.
4. method according to claim 3, is characterized in that, described gas flow is set as that 5 ~ 15SLM, power are 80 ~ 120W.
5. method according to claim 4, is characterized in that, described method also adopts fluid circulation system, and described fluid circulation system adjusts flow velocity by peristaltic pump, and each circular treatment time is 5 ~ 15s; Described microorganism cells suspension is bacterial cell suspension, actinomycetes cells suspension, fungal cell's suspension, alga cells suspension or algae protoplasm somatocyte suspension.
6. the arbitrary described method of claim 1-5 comprises the steps:
1) select the single colony inoculation of microorganism of new activation in liquid nutrient medium from the solid plate of microorganism growth, concussion overnight incubation, obtains bacterium liquid;
2) by step 1) the bacterium liquid that obtains is inoculated in the new liquid nutrient medium changed, and concussion is cultured to bacterium liquid OD
600be 0.3 ~ 1.0;
3) by step 2) concussion gained bacterium liquid adds in sterile centrifugation tube, centrifugal, and abandon supernatant, retain precipitation thalline;
4) with aseptic 0.1mol/LCaCl
2the resuspended step 3 of the aqueous solution) thalline of centrifugation;
5) with described plasma treatment step 4) thalline that obtains;
6) step 5 is collected) thalline after Cement Composite Treated by Plasma, be competent cell.
7. method according to claim 6, is characterized in that, step 2) in, described bacterium liquid OD
600be 0.5 ~ 0.7; The inoculum size of described bacterium liquid is 5% ~ 15%.
8. method according to claim 7, is characterized in that, step 3) in, described is centrifugal, rotating speed 4000 ~ 10000r/min, centrifugation time 0.5 ~ 2.0min.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101624591A (en) * | 2008-07-07 | 2010-01-13 | 清华大学 | Method for breeding microorganisms by plasma-induced mutation |
| CN102321606A (en) * | 2011-08-04 | 2012-01-18 | 大连大学 | Method for enhancing cell membrane permeability by utilizing atmospheric pressure cold plasma |
| CN102382763A (en) * | 2011-12-20 | 2012-03-21 | 北京思清源生物科技有限公司 | Plasma microbial mutagenic breeding equipment |
-
2013
- 2013-12-18 CN CN201310700136.9A patent/CN103710335B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101624591A (en) * | 2008-07-07 | 2010-01-13 | 清华大学 | Method for breeding microorganisms by plasma-induced mutation |
| CN102321606A (en) * | 2011-08-04 | 2012-01-18 | 大连大学 | Method for enhancing cell membrane permeability by utilizing atmospheric pressure cold plasma |
| CN102382763A (en) * | 2011-12-20 | 2012-03-21 | 北京思清源生物科技有限公司 | Plasma microbial mutagenic breeding equipment |
Non-Patent Citations (3)
| Title |
|---|
| 等离子体臭氧对大肠杆菌的杀灭机理研究;顾春英 等;《现代预防医学》;20041231;第31卷(第1期);33-35 * |
| 脉冲放电等离子体灭活铜绿微囊藻的实验研究;王翠华 等;《广东工业大学学报》;20111231;18-21 * |
| 辉光放电等离子体降解有机污染物及杀藻灭菌的研究;夏青;《中国优秀硕士学位论文全文数据库》;20121015;摘要、第3.4.9节 * |
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