CN106520905A - Method for screening environmental stress response promoters - Google Patents
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Abstract
The invention belongs to the technical field of biology and provides a method for screening environmental stress response promoters. The method comprises the steps that a promoter library is established, representation strains of the promoter library are activated and then cultivated under different stress conditions, OD600 and the fluorescence intensities are measured, the fluorescence intensities of different component culture media and under different culture conditions are analyzed, and the environmental response promoters are excavated. The method is based on the natural promoter library of saccharomyces cerevisiae chromosomes V, quantitative presentation and excavation of promoters of a whole chromosome size under specific culture conditions are achieved, and the promoters giving response to different environment conditions are obtained through screening. The method provided by the invention can be further applied to screening of promoters, giving to response to different environment conditions, of other promoter libraries, and screening of promoters giving response to other conditions, and the target stress response promoters can be excavated systematically, conveniently, effectively and visually.
Description
Technical field
The invention belongs to biological technical field, and in particular to a kind of method base of screening environment-stress response promoter should
With.
Background technology
With the further investigation and extensively application of synthetic biology, for various chassis, element and module demand increasingly
Increase, wherein element is the elementary cell for building various functions module and complication system, and the sign and excavation to element is higher
The basis that level through engineering approaches build.Promoter is as conventional gene expression regulation element, according to its different regulation and control type, main
It is divided into two classes：Inducible promoter and constitutive promoter.Constitutive promoter (constitutive promoter) is referred to
Under the control of such promoter, the expression somewhat constant of structural gene on certain level, in different tissues, position expression
Without notable difference.Inducible promoter refer to can controlling gene table is carried out in cell growth process under a certain specified conditions
Reach, the promoter of this type can be significantly increased the transcriptional level of gene.Relative to constitutive promoter, induction type is opened
Mover is capable of achieving efficient, controllable and special regulation and control.At present, existing many inducible promoters are applied to synthetic biology neck
Domain.IPTG (isopropylthiogalactoside) derivable lac, tac and T7 in conventional inducible promoter such as escherichia coli
Promoter, the derivable BAD promoteres of L-arabinose, in saccharomyces cerevisiae, galactose derivable GAL1, GAL7 and GAL10 are opened
Mover, the derivable ADH2 promoteres of ethanol and the derivable CUP1 promoteres of copper ion etc., methanol induction in Pichia sp.
AOX1 promoteres.There are the Light-inducible or temperature inducible promoter of various environment-stress responses in addition.
Traditionally for the condition setting of many function informations based on its related gene of the sign of promoter, it is impossible to excavate latent
It is in unknown conditional response and relatively low to the Efficiency of individual promoter, it is impossible to systematically to characterize in cell global level
Promoter response condition under various environmental stress conditions.In recent years, the research method in cell global level, such as transcription group
It is swift and violent with protein science development, and it is advantageously applied to the gene differential expression that various life entities are responded to varying environment, but its
As a result it is such as the three-dimensional result of genome, the site effect of gene, the sequence of gene by produced by many factors collective effect
Difference in difference, the stability of mRNA, terminator difference, translation skill etc., and regulating effect of the environmental condition to promoter
Simply one of factor.Accordingly, it would be desirable to can with complete system reflect each independent free promoter at different conditions
The method for regulating and controlling identical reporter gene expression.
The content of the invention
In view of this, present invention aims to the diversified demand of environmental response native promoter element and individual
Other promoter characterization is characterized and organizes the deficiency of method, there is provided one kind carries out environmental response startup based on natural promoter library
The method that subsystem is characterized and excavated.
For realizing the purpose of the present invention, the present invention is adopted the following technical scheme that：
A kind of method that screening environment-stress responds promoter, builds promoter library, and activating promoters library characterizes bacterium
Strain, then cultivates under the conditions of Different stress, determines OD600 and fluorescence intensity, to different component culture medium and different culture bars
The fluorescence intensity of part is analyzed, and excavates environmental response promoter.
In some embodiments, the structure promoter library is specially and cuts with reference to S. cerevisiae chromosomal sequence information
Take promoter region；Restriction enzyme site analysis is carried out to each promoter region, designs primer；With saccharomyces cerevisiae genome as template
PCR amplifications obtain target start sub-piece, build restructuring and characterize carrier；Transformed saccharomyces cerevisiae competent cell；Picking monoclonal
Incubated overnight, 25% glycerol are preserved, and form promoter library.
In the method for screening environment-stress response promoter of the present invention, the template is preferably saccharomyces cerevisiae BY4741
Genome.The competent cell is preferably saccharomyces cerevisiae BY4741 competent cells.
Further, the step of also including before the transformed competence colibacillus cell that characterizing carrier to promoter verifies, with
Ensure that promoter characterizes the correct row of carrier.
Preferably, opening of depositing for going bail for of activation described in the method for screening environment-stress of the present invention response promoter
Mover library sign bacterial strain is connected to SC-Leu culture medium and cultivates under the conditions of 30 DEG C, 900rpm.
In some embodiments, the promoter library that the activation is deposited for going bail for characterizes each 5 Μ l of bacterial strain, is connected to 96 holes
The each hole of Tissue Culture Plate, wherein each 200 μ L of hole culture medium containing SC-Leu, the mistake under the conditions of 30 DEG C of micropore plate oscillator, 900rpm
Night cultivates.
Wherein, the SC-Leu culture medium prescriptions are that every 1L contains：20g glucoses, 6.7g yeast nitrogen basis (YNB),
Powder of amino acids of the 2.0g without histidine, uracil, tryptophan and lysine, 10mL2g/L uracil, 10mL2g/L color ammonia
Acid, 10mL 2g/L histidine, it is 6 to adjust pH after constant volume.
In the method for screening environment-stress response promoter of the present invention, culture under the conditions of the Different stress is difference
Carbon source culture medium culturing, high-salt stress culture medium culturing, osmotic pressure stress culture medium culturing, aminoacid defect culture medium culturing
Or high-temperature cultivation.
Wherein, the different carbon source culture medium is the D- galactose with sucrose, and ethanol or glycerol replace glucose as carbon source
SC-Leu culture medium.The high-salt stress culture medium is to add the SC-Leu culture medium culturings of 1M NaCl.The hyperosmosises
Coercing cultivation base is the SC-Leu culture medium for adding 1M Sorbitol.The aminoacid defect culture medium is without powder of amino acids
SC-Leu culture medium or the gala sugar culture-medium without powder of amino acids.
Under above-mentioned stress conditions, culture is 30 DEG C, cultivates under the conditions of 900rpm.I.e. recombinant bacterial strain is transferred in above-mentioned various
In culture medium, more than 12h is cultivated under the conditions of 30 DEG C of micropore plate oscillator, 900rpm.
Described in the method for screening environment-stress response promoter of the present invention, high-temperature cultivation is 39 DEG C, 900rpm conditions
Lower culture.I.e. recombinant bacterial strain is transferred in SC-Leu culture medium, under the conditions of 39 DEG C of micropore plate oscillator, 900rpm cultivates 12h
More than.
In the method for screening environment-stress response promoter of the present invention, before the measure OD600 and fluorescence intensity also
Supernatant is removed including centrifugation, add water resuspended step.The centrifugation is preferably 4000rpm centrifugation 2min.
According to the present invention, the fluorescence intensity under the conditions of Different stress is analyzed specially cultivates to Jing SC-Leu
The promoter that base (blank control group) is cultivated is according to the descending sequence of fluorescence intensity characterization result and draws scatterplot；According to weight
The initiator sequence of row, the fluorescence intensity sequence to each promoter under the conditions of Different stress, draws under varying environment condition stress
The spectrogram of response promoter, screening deviate the promoter of regulation and control normal expression.
If without can present in environment response theory and control medium i.e. SC-Leu culture medium culturings fluorescence intensity sequence phase
Similar smoothed curve figure.But actual under special stressful environmental, some promoteres have response, therefore its fluorescent value can be than in front and back's phase
The promoter of adjacent normal expression is higher, that is, the promoter for deviateing regulation and control normal expression occur.Thus screening obtains environment-stress
The promoter of response.
Present invention also offers described method is applied in screening inducible promoter.
As shown from the above technical solution, the invention provides a kind of method of screening environment-stress response promoter, builds
Promoter library, activating promoters library characterize bacterial strain, then cultivate under the conditions of Different stress, determine OD600 and fluorescence is strong
Degree, is analyzed to the fluorescence intensity of different component culture medium and different condition of culture, excavates environmental response promoter.The present invention
Methods described is based on saccharomyces cerevisiae V chromosome natural promoters library, realizes the promoter of whole chromosome yardstick in specific training
Quantitatively characterizing and excavation under the conditions of supporting, screens the promoter of the varying environment condition that meets with a response.The method of the invention may be used also
The screening of the promoter of other promoter library response varying environment conditions is applied to, also apply be applicable to respond other conditions startup
Target stress response promoter is easily and effectively intuitively excavated in the screening of son, system.The present invention is by the flow process of efficiency standard
Method, can screen the inducible promoter for excavating possible environmental response on genome to greatest extent, meet synthesising biological
The needs that the demand and engineered strain for learning controlling element builds.
Description of the drawings
In order to be illustrated more clearly that the embodiment of the present invention or technical scheme of the prior art, below will be to embodiment or existing
Accompanying drawing to be used needed for having technology description is briefly described.
Fig. 1 shows the V chromosome promoter intensity maps and the figure that reorders of present invention measure；
Fig. 2 shows promoter screening figure under Different stress response condition of culture of the present invention.
Specific embodiment
Below in conjunction with the embodiment of the present invention, the technical scheme in the embodiment of the present invention is clearly and completely described,
Obviously, described embodiment is only a part of embodiment of the invention, rather than the embodiment of whole.Based in the present invention
Embodiment, the every other embodiment obtained under the premise of creative work is not made by those of ordinary skill in the art, all
Belong to the scope of protection of the invention.If no special instructions, reagent involved in the embodiment of the present invention is commercially available prod,
To be obtained by commercial channel purchase.
Wherein Sc-Leu culture medium (per 1L)：20g glucoses, 6.7g yeast nitrogen basis (YNB), 2.0g powder of amino acids
(without histidine, uracil, four kinds of nutrients of tryptophan and lysine), in addition three kinds of 100 × mother solutions (uracil, tryptophans
2g/L is with histidine) each 10mL, it is 6 to adjust pH with NaOH solution after constant volume.
Sucrose medium：In Sc-Leu medium components, 20g/L glucoses replace with 20g/L sucrose.
Gala sugar culture-medium：In Sc-Leu medium components, 20g/L glucoses replace with 20g/L galactose.
Glycerin medium：In Sc-Leu medium components, 20g/L glucoses replace with 20g/L glycerol.
Ethanol culture medium：In Sc-Leu medium components, 20g/L glucoses replace with 20g/L ethanol.
NaCl culture medium：Add final concentration 1M NaCl in Sc-Leu culture medium.
Sorbitol culture medium：Add final concentration 1M Sorbitol in Sc-Leu culture medium.
Amino acid media：Default 2.0g powder of amino acids in Sc-Leu culture medium.
Gala sugar carbon source aminoacid defect culture medium：Default 2.0g powder of amino acids in gala sugar culture-medium.
The culture medium for preparing sterilizes 20 minutes for 121 DEG C in high-pressure steam sterilizing pan.
The structure of 1 promoter library of embodiment.
Saccharomyces Cerevisiae in S 288c V chromosomal DNA sequences, reference sequences are shown in http://www.ncbi.nlm.nih.gov/
Nuccore/NC_001137.3, takes adjacent two protein coding genes intervening sequence in the same direction as promoter region, searches whether to contain
BsaI restriction enzyme sites (GGTCTC or GAGACC) or BsmBI restriction enzyme sites (CGTCTC or GAGACG), contain according to its sequential design
The primer of BsaI or BsmBI restriction enzyme sites.For not only containing BsaI but also containing BsmBI restriction enzyme sites promoter sequence, using ring
Shape polymerase extension method (CPEC) mode builds sign carrier, according to 5 ' and 3 ' each 20bp of its sequence truncation or so and introduces expression
The homologous fragment of carrier is used as primer.
Saccharomyces cerevisiae BY4741 genomes are extracted using phenol/chloroform method, starts sub-pieces by template PCR amplifications of genome
Section.For plasmid vector and the promoter fragment containing BsaI restriction enzyme sites, in 37 DEG C of enzyme action 1 hour；For the position of enzyme action containing BsmBI
The promoter fragment of point, 55 DEG C of enzyme action 1 hour.After enzyme action, carrier segments are connected using T4DNA ligases with promoter fragment, are turned
Change competent escherichia coli cell.CPE C modes are built in sign carrier, and equimolar promoter fragment is passed through with carrier segments
Assemble through ring-type PCR after homologous complementary pairing, subsequently convert competent escherichia coli cell.Converted product even spread card that
Chloramphenicol resistance LB flat boards, 37 DEG C of constant incubator incubated overnight, picking single bacterium colony enter performing PCR checking.Checking is correct to build bacterium
Strain, connects bacterium in 3mL kalamycin resistance LB culture medium, and 37 DEG C, 220rpm incubated overnight, 15% glycerol are preserved, upgrading grain, and are surveyed
Sequence is verified.Characterize carrier to adopt lithium acetate transformation method transformed saccharomyces cerevisiae BY4741 and screen, picking yeast monoclonal is inoculated in
30 DEG C of incubated overnight of SC-Leu fluid mediums, 25% glycerol are preserved, and form promoter library.
Under 2 multiple stress conditions of embodiment, the response of promoter library is characterized
Various Different stress condition used medium difference are as follows：
Different carbon source culture medium is respectively：Sc-Leu culture medium, sucrose medium, gala sugar culture-medium, glycerin medium,
Ethanol culture medium.
High-salt stress conditioned medium：NaCl culture medium.
Osmotic pressure stress conditioned medium：Sorbitol culture medium.
Aminoacid defect culture medium：Sc-Leu aminoacid defect culture medium, gala sugar carbon source aminoacid defect culture medium.
High temperature culture conditions used medium is：Sc-Leu culture medium.
Promoter library seed is activated first, each 5 μ L recombinant Saccharomyces cerevisiae seeds deposited of going bail for, and is connected to 96 hole cell culture
The each hole of plate, wherein each 200 μ L of hole culture medium containing Sc-Leu, the Tissue Culture Plate of sealing is in 30 DEG C of micropore plate oscillator, 900rpm
Under the conditions of incubated overnight.
For other 9 kinds of condition of culture in addition to high temperature culture conditions, the recombinant bacterial strain of incubated overnight is transferred in each of 200 μ L
Culture medium is planted, 12h is cultivated under the conditions of 30 DEG C of micropore plate oscillator, 900rpm, and temperature used by high-temperature cultivation is 39 DEG C.Each group
Tissue Culture Plate under condition of culture is placed in plate centrifuge and 2min is centrifuged in 4000rpm, sucks supernatant, uses per pore fungi body
200 μ L distilled water are resuspended, after concussion is mixed, determine OD in Microplate reader600And green fluorescence intensity, collect different
Data under condition of culture.
3 data processing of embodiment is excavating stress response promoter.
Under 30 DEG C of condition of culture of Sc-Leu culture medium, V chromosome promoter intensity characterization result such as Fig. 1 a of measure
It is shown, it is descending to its result to reorder, and numbering 1-279, as a result as shown in Figure 1 b.
According to the initiator sequence reset, to promoter under other environment-stress condition of culture and intensity sequence.Sucrose is trained
Under foster base condition of culture result as shown in Figure 2 a, under the conditions of galactose culture medium culturing result as shown in Figure 2 b, glycerin medium
Under condition of culture, as shown in Figure 2 c, under the conditions of ethanol culture medium culturing, result is as shown in Figure 2 d for result；Sc-AA culture medium culturing bars
Under part, as shown in Figure 2 e, under the conditions of galactose-AA culture medium culturings, result is as shown in figure 2f for result；NaCl culture medium condition of culture
As shown in Figure 2 g, under the conditions of Sorbitol culture medium culturing, result is as shown in fig. 2h for lower result；Result under 39 DEG C of high temperature culture conditions
As shown in fig. 2i.
By promoter screening figure under each stress response condition of culture, the normal table of deviation regulation and control intuitively can be significantly found
Other promoteres for reaching, in each figures of Fig. 2 at arrow indication, the promoter for as potentially responding to environmental condition, excavation are obtained
Potential saccharomyces cerevisiae V chromosomes environmental response promoter it is as shown in table 1.
1 saccharomyces cerevisiae V chromosome environmental response promoteres of table
Claims (9)
1. a kind of method that screening environment-stress responds promoter, it is characterised in that build promoter library, activating promoters text
Storehouse characterizes bacterial strain, then cultivates under the conditions of Different stress, determines OD600 and fluorescence intensity, to different component culture medium and not
It is analyzed with the fluorescence intensity of condition of culture, excavates environmental response promoter.
2. method according to claim 1, it is characterised in that the structure promoter library is specially and refers to saccharomyces cerevisiae
Chromosome sequence information interception promoter region；Restriction enzyme site analysis is carried out to each promoter region, designs primer；With ferment of making wine
Female genome obtains target start sub-piece for template PCR amplifications, builds restructuring and characterizes carrier；Transformed saccharomyces cerevisiae competence is thin
Born of the same parents；Picking monoclonal incubated overnight, 25% glycerol are preserved, and form promoter library.
3. method according to claim 1, it is characterised in that the activation is that the promoter library deposited of going bail for characterizes bacterial strain
It is connected to SC-Leu culture medium to cultivate under the conditions of 30 DEG C, 900rpm.
4. method according to claim 3, it is characterised in that SC-Leu culture medium prescriptions are that every 1L contains：20g glucoses,
6.7g yeast nitrogen basis (YNB), powder of amino acids of the 2.0g without histidine, uracil, tryptophan and lysine,
10mL2g/L uracil, 10mL2g/L tryptophans, 10mL2g/L histidine, it is 6 to adjust pH after constant volume.
5. method according to claim 1, it is characterised in that the Different stress condition is the training of different carbon source culture medium
Foster, high-salt stress culture medium culturing, osmotic pressure stress culture medium culturing, aminoacid defect culture medium culturing or high-temperature cultivation.
6. method according to claim 1, it is characterised in that the different carbon source culture medium is the D- galactose with sucrose,
Ethanol or glycerol replace glucose as the SC-Leu culture medium of carbon source；The high-salt stress culture medium is addition 1M NaCl
SC-Leu culture medium culturings；The osmotic pressure stress culture medium is the SC-Leu culture medium for adding 1M Sorbitol；The amino
Sour defect culture medium is the SC-Leu culture medium without powder of amino acids or the gala sugar culture-medium without powder of amino acids；
The high-temperature cultivation is 39 DEG C, cultivates under the conditions of 900rpm.
7. method according to claim 1, it is characterised in that also include centrifugation before measure OD600 and fluorescence intensity
Supernatant is removed, add water resuspended step.
8. method according to claim 1, it is characterised in that the fluorescence intensity under the conditions of Different stress is carried out point
Analysis is specially the promoter to Jing SC-Leu culture medium culturings according to the descending sequence of fluorescence intensity characterization result and draws scattered
Point diagram；According to the initiator sequence reset, the fluorescence intensity sequence to each promoter under the conditions of Different stress, screening deviate regulation and control
The promoter of normal expression.
9. the method described in claim 1-8 any one is applied in screening inducible promoter.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110951739A (en) * | 2019-12-25 | 2020-04-03 | 江南大学 | Promoter induced and expressed by high temperature and application thereof |
| CN111254143A (en) * | 2020-01-21 | 2020-06-09 | 天津科技大学 | Construction method of arthrobacter simplex engineering strain with excellent stress tolerance, strain and application thereof |
| CN111733089A (en) * | 2020-05-12 | 2020-10-02 | 北京理工大学 | Strain construction method for improving saccharomyces cerevisiae robustness |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101922047A (en) * | 2009-06-17 | 2010-12-22 | 华东理工大学 | Gap promoter library and application thereof |
| CN105274112A (en) * | 2015-11-20 | 2016-01-27 | 江南大学 | Promoter in induced expression under acidic condition |
| CN105296486A (en) * | 2015-11-18 | 2016-02-03 | 湖北大学 | Promoter library construction method and strong promoter |
| CN105603537A (en) * | 2016-03-11 | 2016-05-25 | 南京工业大学 | Construction method and application of promoter library |
-
2016
- 2016-12-15 CN CN201611162858.3A patent/CN106520905A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101922047A (en) * | 2009-06-17 | 2010-12-22 | 华东理工大学 | Gap promoter library and application thereof |
| CN105296486A (en) * | 2015-11-18 | 2016-02-03 | 湖北大学 | Promoter library construction method and strong promoter |
| CN105274112A (en) * | 2015-11-20 | 2016-01-27 | 江南大学 | Promoter in induced expression under acidic condition |
| CN105603537A (en) * | 2016-03-11 | 2016-05-25 | 南京工业大学 | Construction method and application of promoter library |
Non-Patent Citations (1)
| Title |
|---|
| 易永红: "利用人工启动子文库技术筛选低温敏感性启动子", 《中国优秀硕士学位论文全文数据库》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110951739A (en) * | 2019-12-25 | 2020-04-03 | 江南大学 | Promoter induced and expressed by high temperature and application thereof |
| CN110951739B (en) * | 2019-12-25 | 2021-03-30 | 江南大学 | Promoter induced and expressed by high temperature and application thereof |
| CN111254143A (en) * | 2020-01-21 | 2020-06-09 | 天津科技大学 | Construction method of arthrobacter simplex engineering strain with excellent stress tolerance, strain and application thereof |
| CN111254143B (en) * | 2020-01-21 | 2023-11-03 | 天津科技大学 | Construction method of simple Arthrobacter engineering strain with excellent stress tolerance, strain and application thereof |
| CN111733089A (en) * | 2020-05-12 | 2020-10-02 | 北京理工大学 | Strain construction method for improving saccharomyces cerevisiae robustness |
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