CN103146666B - Streptococcus suis cell wall hydrolase as well as coding gene and application thereof - Google Patents
Streptococcus suis cell wall hydrolase as well as coding gene and application thereof Download PDFInfo
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- CN103146666B CN103146666B CN201310089171.1A CN201310089171A CN103146666B CN 103146666 B CN103146666 B CN 103146666B CN 201310089171 A CN201310089171 A CN 201310089171A CN 103146666 B CN103146666 B CN 103146666B
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Abstract
The invention provides a streptococcus suis cell wall hydrolase as well as a coding gene and an application thereof, and relates to the field of biotechnologies. The amino acid sequence of the streptococcus suis cell wall hydrolase is as shown in SEQ ID NO:1. The invention further discloses the coding gene of the streptococcus suis cell wall hydrolase and a recombinant expression vector and a recombinant bacterium containing the streptococcus suis cell wall hydrolase. The invention provides a method for preparing the streptococcus suis cell wall hydrolase. The streptococcus suis cell wall hydrolase is obtained through cultivating the recombinant bacterium and then purifying. The streptococcus suis cell wall hydrolase can effectively kill streptococcus suis, escherichia coli, staphylococcus aureus and aeromonas hydrophilia. The gene of the streptococcus suis cell wall hydrolase can be efficiently expressed in recombinant escherichia coli, and the activity and stability of the obtained streptococcus suis cell wall hydrolase are high, so that the streptococcus suis cell wall hydrolase can effectively kill streptococcus suis, escherichia coli, staphylococcus aureus and aeromonas hydrophilia.
Description
Technical field
The present invention relates to biological technical field, be specifically related to a kind of swine streptococcus cell wall lytic enzyme, its encoding gene and application.
Technical background
Swine streptococcus is a kind of important Zoonosis disease pathogen, can cause meningitis, sacroiliitis, endocarditis, septicemia, pneumonia, shock and sudden death etc.This bacterium can be divided into 33 serotypes by capsular polysaccharide antigenic difference, wherein 2 types (SS2) are popular the widest, pathogenic the strongest, the cause of disease that swinery carrying rate is higher, to pig industry, cause tremendous economic loss, the life security of public health especially practitioner is also constituted a serious threat.This disease has successively been reported in Holland, Britain, the U.S., Canada, Australia, New Zealand, Belgium, Brazil, Denmark, Norway, Finland, Spain, Germany, Ireland, Japan, TaiWan, China and continent etc.
After Streptococcus suis 2 types infect, generally take antibiotic therapy, but antibiotic abuse in recent years makes streptococcus suis 2-type produce serious resistance, brings difficulty to these sick prevention and control.
Summary of the invention
The object of this invention is to provide a kind of swine streptococcus cell wall lytic enzyme, can effectively kill swine streptococcus.
Another object of the present invention is to provide the gene of swine streptococcus cell wall lytic enzyme.
A further object of the present invention is to provide swine streptococcus cell wall lytic enzyme in the application of killing aspect swine streptococcus.
Object of the present invention adopts following technical scheme to realize:
A swine streptococcus cell wall lytic enzyme, its aminoacid sequence is as shown in SEQ ID NO:1.
The encode gene of described swine streptococcus cell wall lytic enzyme.The sequence of described gene is as shown in SEQ ID NO:2.Be understandable that, due to the degeneracy of amino acid code, the gene of the swine streptococcus cell wall lytic enzyme of encoding in the present invention, except the sequence shown in SEQ ID NO.2, also comprises other nucleic acid molecule of the above-mentioned aminoacid sequence of encoding.
The recombinant expression vector of the gene that contains swine streptococcus cell wall lytic enzyme.The gene order of described swine streptococcus cell wall lytic enzyme can be for shown in SEQ ID NO.2.Described recombinant expression vector is that the gene of swine streptococcus cell wall lytic enzyme is inserted to pET-32a(+) EcoR I and Xho I restriction enzyme site between.
The recombinant bacterium of the gene that contains swine streptococcus cell wall lytic enzyme.The gene order of described swine streptococcus cell wall lytic enzyme can be for shown in SEQ ID NO.2.
A method of preparing swine streptococcus cell wall lytic enzyme, obtains described swine streptococcus cell wall lytic enzyme by after cultivating described recombinant bacterium and carrying out purifying.
Described swine streptococcus cell wall lytic enzyme is in the application of killing aspect swine streptococcus.
Beneficial effect of the present invention:
Swine streptococcus cell wall lytic enzyme provided by the invention, can effectively kill swine streptococcus, intestinal bacteria, streptococcus aureus and have a liking for aqueous vapor pseudomonas bacillus.
The gene of swine streptococcus cell wall lytic enzyme of the present invention can be in recombination bacillus coli high efficient expression, gained swine streptococcus cell wall hydrolytic enzyme activities and stability are high, can effectively kill swine streptococcus, intestinal bacteria, streptococcus aureus and have a liking for aqueous vapor pseudomonas bacillus.
Adopt recombinant bacterium of the present invention to be prepared swine streptococcus cell wall lytic enzyme, method is simple, with low cost.
Accompanying drawing explanation
Fig. 1 is the SDS-PAGE electrophorogram of recombinant expression protein, wherein: 1, the cellular lysate liquid of contrast bacterium; 2, without the cellular lysate liquid of the BL21-pET32a-LytA of IPTG inducing culture; The cellular lysate liquid of the BL21-pET32a-LytA of 3-6, IPTG inducing culture, induction time is respectively 2h, 3h, 4h and 5h; 7, the recombinant expression protein after purifying; M, protein molecular marker.
Fig. 2 is the zymogram lab diagram of checking recombinant expression protein cell wall hydrolytic enzyme activities, wherein: 1, the recombinant expression protein of purifying; 2, contrast bacterium lysate; M, protein molecular marker.
Fig. 3 is the Western-blotting analytical electrophoresis figure of recombinant expression protein, wherein: 1, the cellular lysate liquid of contrast bacterium; 2, the recombinant expression protein after purifying; M, protein molecular marker.
Fig. 4 is the dull and stereotyped bacteriolytic test photo of recombinant expression protein to streptococcus suis 2-type HA9801 strain, what wherein on the scraps of paper 1, drip is the recombinant expression protein of purifying, what on the scraps of paper 2, drip is the BL21 lysate containing empty carrier pET32a through IPTG inducing culture, and what on the scraps of paper 3, drip is PBS damping fluid.
Fig. 5 is the dull and stereotyped bacteriolytic test photo of recombinant expression protein to streptococcus suis 2-type ZY05719 strain, what wherein on the scraps of paper 1, drip is the recombinant expression protein of purifying, what on the scraps of paper 2, drip is the BL21 lysate containing empty carrier pET32a through IPTG inducing culture, and what on the scraps of paper 3, drip is PBS damping fluid.
Fig. 6 is the dull and stereotyped bacteriolytic test photo of recombinant expression protein to streptococcus suis 2-type T15 strain, what wherein on the scraps of paper 1, drip is the recombinant expression protein of purifying, what on the scraps of paper 2, drip is the BL21 lysate containing empty carrier pET32a through IPTG inducing culture, and what on the scraps of paper 3, drip is PBS damping fluid.
Embodiment
Below embodiments of the invention are elaborated, the present embodiment is implemented take technical solution of the present invention under prerequisite, provided detailed embodiment and concrete operating process, the experimental technique of unreceipted actual conditions, principle of work, carry out according to normal condition conventionally.Protection scope of the present invention is not prior to following embodiment.
The public information of streptococcus suis 2-type T15 strain: Vecht, U., J.P.Arends, et al. (1989). " Differences in virulence between two strains of Streptococcus suis type II after experimentally induced infection of newborn germ-free pigs. " Am J Vet Res50 (7): 1037-1043.
The acquisition of embodiment 1 swine streptococcus cell wall lytic enzyme, its encoding gene
By zymogram analysis of experiments the cell wall lytic enzyme of streptococcus suis 2-type T15 strain, find to have a kind of swine streptococcus cell wall lytic enzyme in this bacterial strain, its molecular weight is about 25kDa, dissolved cell wall activity is very strong.
By the full genome analysis to streptococcus suis 2-type T15 strain, find that the sequence cell wall lytic enzyme relevant to streptococcus pyogenes phage as shown in SEQ ID NO:2 exists homology, we are by the sequence called after LytA shown in SEQ ID NO:2.
After streptococcus suis 2-type T15 strain disappearance LytA, its zymogram test is found, loses the dissolving band at 25kDa place, has illustrated that LytA is exactly the encoding gene of swine streptococcus cell wall lytic enzyme.The aminoacid sequence of described swine streptococcus cell wall lytic enzyme is as shown in SEQ ID NO:1.
Although, more than provide from streptococcus suis 2-type T15 strain and obtained LytA,, those skilled in that art, according to gene order disclosed by the invention, adopt additive method, as synthesis method, can access LytA equally, and this is apparent.
The Expression and purification of embodiment 2 swine streptococcus cell wall lytic enzymes:
One. the clone of swine streptococcus cell wall lytic enzyme LytA gene
According to swine streptococcus cell wall hydrolase gene LytA design primer, carry out pcr amplification.Upstream primer LytA-F comprises EcoR I restriction enzyme site, and sequence is GCGGAATTCATGACAACAGTAAATGAAGTAGTTAATTTTGCCAAAG(SEQ IDNO:3).Downstream primer LytA-R comprises Xho I restriction enzyme site, and sequence is GCTCTCGAGCTATTTAAACGTACCATAAGGCACGAC(SEQ ID NO:4).
Pcr amplification system is: the LytA-F1 μ L of 2 * Pfu PCR MasterMix (TIANGEN Biotech (Beijing) Co., Ltd.), 12.5 μ L, 10 μ M/L, the LytA-R1 μ L of 10 μ M/L, streptococcus suis 2-type T15 pnca gene group 0.5 μ L, distilled water 10 μ L.
Pcr amplification program is: 94 ℃ of denaturation 5min; 94 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ are extended 50s, 30 circulations; 72 ℃ are extended 10min
The pcr amplification product of getting 5 μ L detects with 1% agarose gel electrophoresis, and all the other PCR products reclaim test kit (TaKaRa MiniBEST Agarose Gel DNA Extraction Kit) with glue and reclaim.
Two. the structure of recombinant expression vector
With restriction enzyme EcoR I and Xho I(TaKaRa) pcr amplification product is carried out to double digestion, 37 ℃ of incubation 4h, agarose electrophoresis reclaims enzyme and cuts product.
With restriction enzyme EcoR I and Xho I, pET32a carrier is carried out to double digestion, 37 ℃ of incubation 4h, agarose electrophoresis reclaims enzyme and cuts product.
Double digestion reaction system is as follows:
The enzyme that the enzyme of pcr amplification product is cut to product and pET32a carrier is cut T4DNA ligase enzyme for product (TaKaRa) and is carried out ligation (16 ℃ of connections are spent the night).
Ligation system is as follows:
By the evaluation of checking order of connection product, it is pET32a that result shows to have obtained recombinant expression vector pET32a-LytA(skeleton plasmid, between its EcoR I and Xho I restriction enzyme site, has inserted LytA).
Three. the expression of recombinant expression protein
According to ordinary method, recombinant expression plasmid pET32a-LytA is transformed into e. coli bl21 competent cell, after resistance screening (penbritin), random picking list bacterium colony, be inoculated in containing in the LB liquid nutrient medium of penbritin 100 μ g/mL, 37 ℃ of shaking culture 6～8h, directly get bacterium liquid and carry out PCR reaction, preliminary screening positive colony.Through PCR, be accredited as positive clone's bacterium colony, amplification cultivation, extracting plasmid is also identified by double digestion, is obtained positive recombinant bacterium BL21-pET32a-LytA.
By the overnight culture of positive recombinant bacterium BL21-pET32a-LytA in 1:100(volume ratio) ratio be inoculated in (containing penbritin 100 μ g/mL) in LB liquid nutrient medium, 37 ℃ of shaking culture.When OD600 value is 0.4～0.6, add IPTG to final concentration 1mM/L, 37 ℃, 180rpm continue shaking culture 2h, 3h, 4h and 5h.Carry out following experiment: IPTG inducing culture, containing the BL21 of empty carrier pET32a, does not add IPTG without IPTG inducing culture BL21-pET32a-LytA(in culturing process) simultaneously.
The lysate of following each bacterium of preparation: the BL21-pET32a-LytA(induction time of IPTG inducing culture is for being respectively 2h, 3h, 4h and 5h), without the BL21-pET32a-LytA of IPTG inducing culture and the BL21(containing empty carrier pET32a of IPTG inducing culture contrast bacterium, lower with).The method of preparing lysate is: by thalline ultrasonication, after centrifugal, get supernatant liquor, this supernatant liquor is lysate.
SDS-PAGE electrophoresis detection: in order to study the expression of recombinant bacterium target protein, the lysate of each thalline is carried out to SDS-PAGE electrophoresis detection.The separation gel of preparation 12% and 5% concentrated glue, boiling water boiling 5min after sample is mixed with sample-loading buffer, carries out electrophoresis: concentrated glue voltage stabilizing 80V, separation gel voltage 120V, electrophoresis 2h under the following conditions.After electrophoresis finishes, with coomassie brilliant blue staining, spend the night, decolouring is observed.Result, as Fig. 1, can see that obvious protein band appears in the lysate of the BL21-pET32a-LytA of inducing culture at 43kD place, recombinant expression protein successful expression is described, and expresses with soluble form.
The purifying of recombinant expression protein: positive recombinant bacterium BL21-pET32a-LytA is seeded to (containing penbritin 100 μ g/mL) in LB liquid nutrient medium, 37 ℃ of shaking culture.Work as OD
600value is 0.4～0.6 o'clock, adds IPTG to final concentration 0.6mM/L, 37 ℃, 180rpm continuation shaking culture 4h.Collect culture, ultrasonication, centrifugal rear collection supernatant are as crude enzyme liquid.Crude enzyme liquid is filtered with the filter membrane of 0.45 μ m, then use HisTrap
tMhP Columns(GE Healthcare) (1mL) purifying, obtains the recombinant expression protein after purifying.Recombinant expression protein after purifying is carried out to SDS-PAGE electrophoresis detection (the results are shown in Figure 1).Can see, crude enzyme liquid is through HisTrap
tMafter the purifying of HP Columns, removed a lot of foreign proteins, on SDS-PAGE electrophoretogram there is obvious protein band in 43kD place.With the antibody obtaining after the recombinant expression protein immunity new zealand rabbit after purifying, as primary antibodie, in order in Western-blotting immunoblotting, use.
Adopt streptococcus suis 2-type bacterium HA9801 cell walls to carry out zymogram test, analyze the cell wall lytic enzyme hydrolysis properties of recombinant expression protein, take and contrast bacterium (BL21 containing empty carrier pET32a of IPTG inducing culture) lysate for contrasting.Method is as follows: join 12% separation gel, streptococcus suis 2-type bacterium HA9801 cell walls is added in SDS-PAGE glue in 0.1% (wt/vol) ratio, state is translucent after gelling is solid.After testing sample is mixed with sample-loading buffer, boil 3min, get 12 μ l and add in loading hole and start electrophoresis, electrophoresis step is identical with SDS-PAGE.Electrophoresis is complete, and glue is soaked in 200ml deionized water, and room temperature vibration 30min, changes deionized water and soak 30min again.SDS-PAGE glue is transferred to (0.1%Triton X-100,10mM MgCl in 200ml renaturation buffer
2, 25mM Tris-HCl; PH7.5), 37 ℃ are shaken 12h slowly, make lytic enzyme renaturation.Whether the separation gel of observing interpolation cell walls has transparent strip to occur.
As shown in Figure 2, recombinant expression protein swimming lane has and very significantly dissolves band at 43kDa place result, illustrates that recombinant expression protein can produce very significantly dissolved cell wall effect; Recombinant expression protein swimming lane exists many to dissolve band at 25-35kDa place, may be because recombinant expression protein be degraded, and the degraded product of generation still has hydrolytic activity; Contrast is not dissolved band and is produced.Zymogram experiment confirms that recombinant expression protein has the activity of dissolving streptococcus suis 2-type cell walls, and this albumen is cell wall lytic enzyme.
Western-blotting immunoblotting: by the gel after above-mentioned SDS-PAGE electrophoresis, PVDF(polyvinylidene difluoride) film and filter paper balance 30min in Tris-glycine transferring film damping fluid.In transfer groove, from anode to negative electrode, stack according to the order of sequence: filter paper, pvdf membrane, gel, filter paper, electric current setting is calculated according to the area of glue: 0.8mA/cm2 calculates, transfer printing 2h.After transferring film finishes, pvdf membrane is put in confining liquid (containing the TBS of 5% skimming milk, 0.5% tween 20, lower same), 4 ℃ are spent the night.Abandon confining liquid, with washings (containing the TBS of 0.5% tween 20, lower same) washing 3 times, each 5min.Add the primary antibodie with confining liquid dilution, under 37 ℃, 60rpm condition, hatch 1.5h.With washings, wash 3 times each 5min.Pvdf membrane is added to the green skies of goat anti-rabbit igg-HRP(biotechnology research institute) in working fluid, under 37 ℃, 60rpm condition, hatch 1h.With washings, wash (5min) 1 time, with Tris-Cl damping fluid (pH7.5), wash 3 times, each 5min.After HRP-DAB substrate colouring reagents box (TIANGEN Biotech (Beijing) Co., Ltd.) colour developing, take pictures, see Fig. 3.As can see from Figure 3, there is obvious band in the recombinant expression protein after purifying, illustrates that this albumen has obvious immunogenicity at 43kD place, can stimulate body to produce antibody.
The external bacteriolyze spectrum of embodiment 3 swine streptococcus cell wall lytic enzymes
Adopt dull and stereotyped bacteriolytic test to detect the external bacteriolyze spectrum of the recombinant expression protein (embodiment 2 obtains) after purifying, concrete grammar is as follows: at Todd-Hewitt broth (THB, Oxoid) the streptococcus suis 2-type bacterium of coating fresh culture on flat board: HA9801, ZY05719 and T15 strain, by the coating of bacterium liquid evenly.After absorbing completely, put into three through autoclaved circular paper, on circular paper, drip respectively the recombinant expression protein after purifying, lysate and the PBS damping fluid of contrast bacterium.Flat board is put to 37 ℃ of diffusion 18h, observed and recorded bacteriolyze situation.Dull and stereotyped bacteriolytic test is found: on circular paper, dripped after the recombinant expression protein after purifying, these scraps of paper have produced obvious transparent bacteriolyze circle around, and dripping the contrast lysate of bacterium and the scraps of paper of PBS damping fluid does not have bacteriolysis around, sees Fig. 4, Fig. 5 and Fig. 6.Dull and stereotyped bacteriolytic test presentation of results: recombinant expression protein is dissolution of bacteria produce bacteriolyze circle significantly effectively, provides new antibiotic preparation for treating the infection that streptococcus suis 2-type causes.
Adopt reduced turbidity method to measure the bacteriolyze spectrum of recombinant expression protein.By strain culturing to be checked to logarithmic phase (OD
600for 0.6-0.8), centrifugal acquisition bacterial sediment.By precipitation with PBS damping fluid wash twice rear resuspended, adjust bacterial turbidity OD
600value is 1.0, adds the recombinant expression protein after 100 μ g purifying, and 37 ℃ of water-bath effect 15min, record OD
600data.Calculate each bacterial turbidity OD
600the drop-out value of value, the results are shown in Table 1.Streptococcus suis 2-type bacterial strain is very responsive to the bacteriolysis of recombinant expression protein, and swine streptococcus 8 types and 9 type bacterial strain susceptibility and 2 types approach, but 7,17,18 types completely can not reorganized expressing protein cracking.Recombinant expression protein is to the gram-positive bacteria of non-swine streptococcus as streptococcus aureus, and streptococcus agalactiae can slight cracking, and to gram-negative bacteria, as intestinal bacteria, Aeromonas hydrophila also can present solute effect to a certain degree.
Table 1 reduced turbidity method measurement result
SEQUENCE LISTING
<110> Agricultural University Of Nanjing
<120> swine streptococcus cell wall lytic enzyme, its encoding gene and application
<130> 20130319
<160> 4
<170> PatentIn version 3.3
<210> 1
<211> 239
<212> PRT
<213> streptococcus suis 2-type T15 strain
<400> 1
Met Thr Thr Val Asn Glu Val Val Asn Phe Ala Lys Asp Leu Ala Asn
1 5 10 15
Arg Gly Gln Gly Val Asp Tyr Asp Gly Trp Tyr Gly Asn Gln Cys Val
20 25 30
Asp Leu Pro Asn Trp Ile Cys Gly Lys Phe Phe Gly Lys Pro Leu Trp
35 40 45
Gly Asn Ala Ile Asp Leu Ile Lys Ser Ala Lys Gln His Gly Phe Glu
50 55 60
Val His Tyr Met Pro Thr Ser Glu Arg Pro Arg Pro Gly Ala Ile Phe
65 70 75 80
Val Lys Asn Tyr Trp Ala Ser Asp Gly Val Asn Tyr Gly His Thr Gly
85 90 95
Leu Ile Ile Gly Val Ser Gly Asn Ala Val Gln Thr Ile Glu Gln Asn
100 105 110
Leu Val Gly Asn Leu Ser Val Gly Gly Pro Ala Gln Tyr Ala Ser Gln
115 120 125
Gln Ile Ser Asn Leu Val Gly Trp Phe Tyr Pro Pro Tyr Ser Asp Ser
130 135 140
Thr Ala Val Ala Thr Gln Ala Ser Ser Gly Asn Leu Gly Lys Tyr Lys
145 150 155 160
Asp Glu Gln Gly Thr Met Thr Val Lys Val Ser Leu Leu Asn Val Arg
165 170 175
Asp Lys Pro Ser Leu Asp Gly Lys Ile Val Ala Thr Tyr Thr Tyr Ser
180 185 190
Glu Gln Phe Asn Tyr Asp Ser Ile Tyr Ile Ala Asp Gly Tyr Phe Trp
195 200 205
Val Ser Tyr Val Ser Arg Ser Gly Val Arg Arg Tyr Val Ala Ala Gly
210 215 220
Glu Glu Ser Asn Arg Arg Asn Val Val Pro Tyr Gly Thr Phe Lys
225 230 235
<210> 2
<211> 720
<212> DNA
<213> streptococcus suis 2-type T15 strain
<400> 2
atgacaacag taaatgaagt agttaatttt gccaaagacc tagccaaccg tggacaaggt 60
gtagactatg atggttggta cggtaatcaa tgtgtagact tgcctaactg gatttgcgga 120
aaattcttcg gcaagccttt gtggggcaat gccattgatt tgataaagtc agccaagcaa 180
cacggctttg aggtgcatta catgcctact tcagaacgtc cacgtccagg ggctatcttt 240
gtcaagaatt actgggccag tgatggtgtc aactatgggc atacgggttt gattatcggt 300
gttagtggca atgcagtcca aaccattgag caaaatcttg ttggtaatct gtctgtcggt 360
ggtcctgctc aatatgctag ccagcaaatc agcaatcttg ttggttggtt ctatccacct 420
tacagcgact ctactgcggt ggcaacacag gcaagcagtg gcaaccttgg aaaatacaaa 480
gacgagcaag ggacaatgac tgttaaagtg tctttgctca atgttcgaga caagcctagt 540
cttgacggta aaattgtggc tacttacact tatagcgagc aatttaacta tgattcgatc 600
tatattgctg acggttactt ttgggtatcg tacgttagcc gtagtggtgt gcgtcgctat 660
gtagcagccg gcgaggagtc aaatcgacgc aatgtcgtgc cttatggtac gtttaaatag 720
<210> 3
<211> 46
<212> DNA
<213> artificial
<220>
<223> upstream primer LytA-F
<400> 3
gcggaattca tgacaacagt aaatgaagta gttaattttg ccaaag 46
<210> 4
<211> 36
<212> DNA
<213> artificial
<220>
<223> downstream primer LytA-R
<400> 4
gctctcgagc tatttaaacg taccataagg cacgac 36
Claims (9)
1. a swine streptococcus cell wall lytic enzyme, its aminoacid sequence is as shown in SEQ ID NO:1.
2. the gene of swine streptococcus cell wall lytic enzyme described in the claim 1 of encoding.
3. gene according to claim 2, is characterized in that: the sequence of described gene is as shown in SEQ ID NO:2.
4. the recombinant expression vector that contains gene described in claim 2 or 3.
5. recombinant expression vector according to claim 4, is characterized in that this recombinant expression vector is that the gene of swine streptococcus cell wall lytic enzyme is inserted to pET-32a's
ecor I and
xhobetween I restriction enzyme site.
6. the recombinant bacterium that contains gene described in claim 2 or 3.
7. recombinant bacterium according to claim 6, is characterized in that: described recombinant bacterium is that recombinant expression vector described in claim 5 is transformed to e. coli bl21 gained.
8. a method of preparing swine streptococcus cell wall lytic enzyme, is characterized in that: by recombinant bacterium described in cultivation claim 6 or 7 and after carrying out purifying, obtain described swine streptococcus cell wall lytic enzyme.
9. described in claim 1, swine streptococcus cell wall lytic enzyme is killed the application aspect the antibiotic preparation of swine streptococcus in preparation.
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| CN102198265A (en) * | 2011-03-22 | 2011-09-28 | 上海交通大学 | Method for degrading streptococcus suis biofilm by applying phage lyase |
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| CN102198265A (en) * | 2011-03-22 | 2011-09-28 | 上海交通大学 | Method for degrading streptococcus suis biofilm by applying phage lyase |
Non-Patent Citations (5)
| Title |
|---|
| Tang,F.et al..AGF87355.1：putative lysin [Streptococcus phage phi30c].《GenBank》.2013, * |
| 噬菌体裂解酶的抗菌特性;王琰等;《微生物学报》;20091004;1277-1281 * |
| 猪链球菌2型新的感染相关因子自溶素的鉴定与分析;顾宏伟等;《微生物学报》;20080104;68-72 * |
| 王琰等.噬菌体裂解酶的抗菌特性.《微生物学报》.2009,1277-1281. |
| 顾宏伟等.猪链球菌2型新的感染相关因子自溶素的鉴定与分析.《微生物学报》.2008,68-72. |
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