CN102993296A - Bovine lactoferricin and preparation method thereof - Google Patents
Bovine lactoferricin and preparation method thereof Download PDFInfo
- Publication number
- CN102993296A CN102993296A CN2011102721788A CN201110272178A CN102993296A CN 102993296 A CN102993296 A CN 102993296A CN 2011102721788 A CN2011102721788 A CN 2011102721788A CN 201110272178 A CN201110272178 A CN 201110272178A CN 102993296 A CN102993296 A CN 102993296A
- Authority
- CN
- China
- Prior art keywords
- aga
- gct
- aag
- ggt
- gtt
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Peptides Or Proteins (AREA)
Abstract
The invention relates to bovine lactoferricin and a preparation method thereof. Bovine lactoferricin (Lfcin B) is 25 amino acid residues hydrolyzed from a bovine lactoferrin (BLF) N-terminal (17-41) by pepsin, and has a molecular weight of approximately 3.1kd. The preparation method comprises the steps of: (2) recombinant expression vector construction; (2) fusion protein expression induction; and (3) protein separation and purification. Bovine lactoferricin has the advantages that: Lfcin B has the highest activity among all the Lfcins, and an antibacterial effect 400 times that of bovine lactoferrin, such that Lfcin B can play an important role in gastrointestinal tracts.
Description
Technical field
The invention belongs to biological technical field, be specifically related to Bovinelactoferrin peptide and preparation method thereof.
Background technology
Microbiotic uses in animal-feed as antibiotic and growth-promoting additive very long history, but the raising along with people's level of understanding, microbiotic being widely used in animal-feed exposes increasing negative effect, wherein serious, the one, can cause the drug residue in the animal products, directly affect the healthy of human consumer; The 2nd, have a strong impact on the microecological balance of animal digestive tract, cause the generation of some pathogenic bacteria resistance to drugs, so that become very difficult for prevention and the treatment by the caused infection of these pathogenic bacterias.At present, resistance problem ubiquity, and the quantity of Resistant strain also significantly increases, some threatened human life's disease originally, such as pulmonary tuberculosis etc., began heavily again to occur on a large scale.Denmark forbids using antibiotic country the earliest in feed, since 1994, gradually reduces the use of microbiotic in feed.2003, the World Health Organization (WHO) suggestion gradually reduced antibiotic use according to the pattern of Denmark.2006, European Union completely forbade the use of microbiotic in animal-feed.Do not stop microbiotic as the use of fodder additives although China also makes laws, gradually reducing usage quantity yet.But China still has more than 20 kind microbiotic to use in animal-feed at present.Along with China joined WTO, antibiotic use has had a strong impact on China's livestock industry product competitive power in the international market.Therefore, the development of new Substitutes For Antibiotic to China's livestock industry safe, healthy, harmonious development is more and more important.
Antibacterial peptide (antibacterial peptides) refers to be present in the small molecule polypeptide that has the extraneous pathogenic micro-organism infringement of opposing in the organism, eliminates the vivo mutations cell, and it is an important component part of animal immune system of defense.Antibacterial peptide is by biological specific gene coding, has broad spectrum antibiotic activity, especially Resistant strain is had obvious killing action, and does not destroy biological cell, non-immunogenicity.Antibacterial peptide can roughly be divided into following a few class: the Magainin (magainin) of cecropin class (cecropins), proline rich residue, the melittin (melittin) that is rich in glycine, the alexin (defensin) that is rich in cystine residue and proline rich and arginic cathelicidin family antibacterial peptide class.Antibacterial peptide has Effective Anti bacterium, fungi, virus, even anti-tumor activity, all has broad application prospects on agricultural and clinical treatment, is the Substitutes For Antibiotic of tool DEVELOPMENT PROSPECT
Summary of the invention
The purpose of this invention is to provide a kind of Bovinelactoferrin peptide (Bovine Lactoferricin, be abbreviated as LfcinB) and preparation method thereof, Lfcin B is active the highest among all Lfcin, and fungistatic effect is 400 times of Bovinelactoferrins, can bring into play prior effect in gi tract.
The objective of the invention is to be achieved through the following technical solutions:
The nucleotides sequence of Bovinelactoferrin peptide is classified as:
GCT?CCA?AGA?AAG?AAC?GTT?AGA?TGG?TGT?ACT?ATT?TCT?CAA?CCA?GAA
TGG?TTT?AAG?TGT?AGA?AGA?TGG?CAA?TGG?AGA?ATG?AAG?AAG?TTG?GGT
GCT?CCA?TCT?ATT?ACT?TGT?GTT?AGA?AGA?GCT?TTC?GCT?TTG?GAA?TGT
ATT?AGA?GCT?ATT?GCT?GAG?AAG?AAG?GCT?GAT?CCT?GTT?ACT?TTG?GAT
GGT?GGT?ATG?GTT?TTC?GAA?GCT?GGT?AGA?GAT?CCA?TAC?AAG?TTG?AGA
CCA?GTT?GCT?GCT?GAA?ATT?TAC?GGT?ACT?AAG?GAA?TCT?CCA?CAA?ACT
CAT?TAC?TAC?GCT?GTT?GCT?GTT?GTT?AAG?AAG?GGT?TCT?AAC?TTT?CAA
TTG?GAT?CAA?TTG?CAA?GGT?AGA?AAG?TCT?TGT?CAT?ACT?GGT?TTG?GGT
AGA?TT
Wherein the active centre of Bovinelactoferrin peptide nucleotide sequence is:
TTT?AAG?TGT?AGA?AGA?TGG?CAA?TGG?AGA?ATG?AAG?AAG?TTG?GGT?GCT
CCA?TCT?ATT?ACT?TGT?GTT?AGA?AGA?GCT?TTC
The aminoacid sequence of this Bovinelactoferrin peptide is:
aprknvrwct?isqpewfkcr?rwqwrmkklg?apsitcvrra?faleciraia?ekkadavtld?ggmvfeagrdpyklrpvaae?iygtkespqt?hyyavavvkk?gsnfqldqlq?grkschtglgr
Wherein the active part of Bovinelactoferrin peptide ammino acid sequence is:
fkcrrwqwrm?kklgapsitc?vrraf。
This sequence is altogether by 365 based compositions, wherein, 5 ' end signal peptide comprises 48 bases, 3 ' end fusion rotein partly comprises 240 bases, the active centre is comprised of 75 bases (from the 49th to the 123rd), coding has the Bovine Lactoferricin albumen of the aminoacid sequence in the sequence table, in the coding region, A accounts for 28.1% (102), C accounts for 14.3% (52), and G accounts for 26.2% (95), and T accounts for 31.4% (114), A+T accounts for 59.5% (216), and C+G accounts for 40.5% (147).
The preparation method of Bovinelactoferrin peptide may further comprise the steps:
1, the structure of recombinant expression vector
1.1, Bovinelactoferrin peptide gene synthetic;
Front 1～121 aminoacid sequence of synthetic Bovinelactoferrin.
1.2, Bovinelactoferrin peptide gene cloning;
The synthetic pair of primers, and add respectively BamH I and Xho I restriction enzyme site at its 5 ' end.Upstream primer and downstream primer are as follows:
Blf-F：5′-TATC
GGATCCGCTCCAAGAAAGAACGTT-3′，
Blf-R：5′-GGTG
CTCGAGTTATCTACCCAAACCAGTATG-3′。
1.3, make up recombinant expression vector.
2, abduction delivering fusion rotein
2.1, contain the acquisition of the engineering bacteria of Bovinelactoferrin peptide gene;
2.2, the abduction delivering fusion rotein.
3, the isolation and purification of protein
3.1, the separation of fusion rotein;
3.2, fusion rotein concentrated;
3.3, the purifying of fusion rotein;
3.4, the preservation of Bovinelactoferrin peptide.
Beneficial effect of the present invention is:
1, two reverse β-pleated sheet structure structures that in the aqueous solution, are amphipathic, have heat-resisting, be difficult for being degraded at digestive tube, the feature of no antigen, can regulate immunization, have broad-spectrum antibacterial action, do not disturb the characteristics such as intestinal beneficial bacterium;
2, it all has anti-microbial effect to many G+ and G-, such as intestinal bacteria, Salmonella enteritidis, Pseudomonas aeruginosa, streptococcus aureus, clostridium perfringens etc., but do not act on bifidus bacillus etc. to the human body beneficial bacterium, simultaneously can also irritation cell growth, participate in immunomodulatory;
3, because its special function as the development research of fodder additives with food preservatives, has been subjected to the attention of domestic and international research institution and manufacturers, become study hotspot.
Embodiment
The embodiment genetically engineered is produced the Bovinelactoferrin peptide
1, the structure of recombinant expression vector
1.1, Bovine Lactoferricin I gene synthetic
Front 1～121 aminoacid sequence of synthetic Bovinelactoferrin: (underscore partly is Bovinelactoferrin peptide (being total to 363bp, 121 amino acid)
GCT?CCA?AGA?AAG?AAC?GTT?AGA?TGG?TGT?ACT?ATT?TCT?CAA?CCA?GAA
TGG?
TTT?AAG?TGT?AGA?AGA?TGG?CAA?TGG?AGA?ATG?AAG?AAG?TTG?GGT
GCT?CCA?TCT?ATT?ACT?TGT?GTT?AGA?AGA?GCT?TTC?GCT?TTG?GAA?TGT
ATT?AGA?GCT?ATT?GCT?GAG?AAG?AAG?GCT?GAT?GCT?GTT?ACT?TTG?GAT
GGT?GGT?ATG?GTT?TT℃GAA?GCT?GGT?AGA?GAT?CCA?TAC?AAG?TTG?AGA
CCA?GTT?GCT?GCT?GAA?ATT?TAC?GGT?ACT?AAG?GAA?TCT?CCA?CAA?ACT
CAT?TAC?TAC?GCT?GTT?GCT?GTT?GTT?AAG?AAG?GGT?TCT?AAC?TTT?CAA
TTG?GAT?CAA?TTG?CAA?GGT?AGA?AAG?TCT?TGT?CAT?ACT?GGT?TTG?GGT
AGA?TT
Be stored among the pUC19/TOP10.
1.2, Bovine Lactoferricin I gene cloning
The synthetic pair of primers, and add respectively BamH I and Xho I restriction enzyme site at its 5 ' end.Upstream primer and downstream primer are as follows:
Blf-F：5′-TATC
GGATCCGCTCCAAGAAAGAACGTT-3′，
Blf-R：5′-GGTG
CTCGAGTTATCTACCCAAACCAGTATG-3′。
Take above-mentioned Blf-F and Blf-R as primer, carry out pcr amplification take front 1～121 aminoacid sequence of synthetic Bovinelactoferrin as template; Amplification system is: 10 * buffer, 2.5 μ L, dNTPmix (10mM) 2 μ L, MgCl
2(25mM) 1.5 μ L, Blf-F primer (10 μ M) 1 μ L, Blf-R primer (10 μ M) 1 μ L, TaKaRa pfu-Taq (5u) 0.25 μ L, template 25ng complements to 25.0 μ L with the sterilization distilled water; Amplification program is: 94 ℃ of denaturation 5min, and 94 ℃ of sex change 30s, 58 ℃ of annealing 30s, 72 ℃ are extended 40s, and totally 35 circulations are at last at 72 ℃ of downward-extension 10min.
Pcr amplification obtains the fragment of 378bp, the PCR product that obtains is reclaimed, connect pMD18-T cloning vector (available from precious biotechnology (Dalian) company limited), obtain recombinant vectors pMD18-T-Bovine Lactoferricin I, then this recombinant vectors is transformed intestinal bacteria, then screening positive clone is identified with BamH I and Xho I double digestion positive colony, send the order-checking of Invitrogen company after evaluation is correct.
1.3, make up recombinant expression vector
The pMD18-T-Bovine Lactoferricin I that above-mentioned steps 1.2 is obtained is with BamH I and Xho I double digestion, the fragment that contains Bovine Lactoferricin I gene that obtains is connected with pET30a (+) expression vector of the same double digestion of warp, obtains pET30a (+)-Bovine Lactoferricin I.
2, abduction delivering fusion rotein
2.1, contain the acquisition of the engineering bacteria of Bovinelactoferrin peptide gene
1) with electric shocking method the recombinant expression vector pET30a (+) of step 1.3 in the step 1-Bovine Lactoferricin I is transformed in intestinal bacteria Rosetta (DE3) competent cell (being purchased from Merck KGaA company).
Wherein the step of electric shocking method is as follows: take out intestinal bacteria Rosetta (DE3) competent cell of-70 ℃ of preservations, ice bath melts; Get Rosetta (DE3) competent cell of 80ul, add the recombinant expression vector that has connected, mixing in the ice-cold micro-aseptic centrifuge tube of 0.5ml, place on ice 5min (competent cell is fully contacted with plasmid), then rapidly mixing liquid is added sterilized clean electricity and transform in the cup, 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 electric shock instrument into, by the parameter of setting, starts the electricimpulse to cell.The LB nutrient solution that has shocked by electricity and added fast immediately 800ul during then sucking liquid is packed the 1.5ml centrifuge tube into, is put into 37 ℃, 150r/min shaking table cultivation 1.5h.(contain penbritin, its final concentration is 50～100ug/ml to the cultivation bacterium liquid bed board of absorption 50ul; Contain paraxin, its final concentration is the LB culture plate of 34ug/ml), 37 ℃ of overnight incubation 14h (half an hour is put in first front, allows nutrient solution be absorbed fully by agar, again the reverse side overnight incubation).
2) screening of positive transformant and evaluation: the mono-clonal bacterium colony of picking 5-8 tool penbritin and 2 kinds of resistances of paraxin from the LB flat board, be inoculated in the 3ml LB liquid nutrient medium that contains 50-100mg/L penbritin and 34mg/L paraxin 37 ℃ of shaking culture 14h of 250rpm.Getting 1-2 μ l bacterium liquid is template, and the Blf-F in the step 1 and Blf-R carry out pcr amplification as primer, whether contain the goal gene fragment in the identification of escherichia coli, pcr amplification reaction condition and thermal cycling arrange with step 1, and the result shows, contains the goal gene fragment in this bacterial strain.Then extract the plasmid in this bacterial strain, with BamH I and Xho I it being carried out double digestion detects, the result shows and contains in the enzyme slitting band and amplification gene band of the same size, illustrates that pET30a (+)-Bovine Lactoferricin I recombinant expression vector successfully changes in intestinal bacteria Rosetta (DE3) cell.
2.2, the abduction delivering fusion rotein
(contain penbritin, its final concentration is 50～100ug/ml in the LB of 20ml nutrient solution to get the positive monoclonal inoculation; Contain paraxin, its final concentration is 34ug/ml) in, 37 ℃, 250r/min shaking table incubated overnight 14～16h.Express in new LB liquid nutrient medium by 1% inoculation bacterium liquid: namely draw 200ul bacterium liquid, (contain penbritin, its final concentration is 50～100ug/ml to be inoculated in the new LB nutrient solution of 20ml; Contain paraxin, its final concentration is 34ug/ml) in, 37 ℃, the about 2h of 250r/min shaking table cultivation are to OD
600=0.4～0.6 o'clock, each sample was drawn 1.0ml in 4 ℃ of preservations, contrasts as initial sample; Then add IPTG and carry out abduction delivering.Add IPTG and make its final concentration reach 0.4mg/ml, then nutrient solution is placed 16 ℃, the cultivation of 250r/min shaking table, collect bacterium liquid in the 50ml centrifuge tube respectively at 16-18h.4 ℃, 8000xg, centrifugal 10min abandons supernatant, adds the complete resuspended precipitation of PBS of 1.0ml precooling (4 ℃), the washing thalline; 4 ℃, 8000xg, centrifugal 10min abandons supernatant, collects somatic cells, claims weight in wet base; Ratio in 1g: 5ml adds the resuspended precipitation of pure water ,-80 ℃, freezes 15min; 100 ℃, boiling 5min repeatedly freezes and adds 5 * Loading Buffer after boiling 2～3 times, behind the centrifugal 3min of maximum speed of revolution, get supernatant and carry out the SDS-PAGE detection, the protein electrophorese result shows the fusion rotein great expression in intestinal bacteria Rosetta (DE3) that contains the Bovinelactoferrin peptide, and its expression amount accounts for 35% of tropina total amount.
3, the isolation and purification of protein
3.1, the separation of fusion rotein
Until 37 ℃ of inducing culture of thalline after 3 hours, 4 ℃, the centrifugal collection thalline of 8000 * g.Behind physiological saline washing thalline, 4 ℃, the centrifugal collection thalline of 8000 * g claims the thalline weight in wet base.Then the ratio in thalline weight in wet base 1g: 10ml adds the resuspended thalline of lysate.The ultrasonic disruption cell, behind DNase I and RNase A enzyme processing nucleic acid, 4 ℃, 10000 * g is centrifugal, collects supernatant liquor, and soluble fusion protein namely is present in the supernatant liquor.
3.2, fusion rotein concentrated
During with 0 ℃, saturation ratio is fusion rotein in the concentrated supernatant of 65% ammonium sulfate, and making its concentration ratio is 10: 1.Then measure protein concn in the concentrated solution with the Bradford method.
3.3, the purifying of fusion rotein
Because fusion rotein contains the His-tag label protein at the N end, so the first step Ni
2+-IMAC affinity chromatography purified fusion protein.Used filler is the IMAC polyacrylamide medium of Bio-Rad company, and the chromatography column specification is 1.5mm * 10cm, and used filler is 10ml in chromatography process, and its dynamic carrying capacity is 4mg albumen/ml filler, and the chromatography Peak Flow Rate is 3ml/min.
Collect elution peak, then regulate the pH value to 2.0 of elutriant, be that 3% amount added stomach en-Pepsin 1: 10000 in mass ratio in elutriant, hatched 1 hour for 37 ℃, make it fully cut label protein and reduce natural lactoferricin, 72 ℃ of thermal treatment 15min then, termination reaction,, conditioned reaction liquid PH to 7.0.
Because lactoferricin belongs to small-molecular peptides, molecular weight is 3.1KDa, and the molecular weight of fusion tag albumen is 10KDa, so the difference of both molecular weight adopts natural peptide fragment and fusion tag after the gel-filtration chromatography separation cuts, collect elution peak, collect lactoferricin.
3.4, the preservation of lactoferricin
Elutriant is carried out lyophilize process after dialysis treatment, collect the lactoferricin lyophilized powder, in 4 ℃ of prolonged preservation.
Claims (6)
1. a Bovinelactoferrin peptide is characterized in that, the nucleotides sequence of this Bovinelactoferrin peptide is classified as:
GCT?CCA?AGA?AAG?AAC?GTT?AGA?TGG?TGT?ACT?ATT?TCT?CAA?CCA?GAA
TGG?TTT?AAG?TGT?AGA?AGA?TGG?CAA?TGG?AGA?ATG?AAG?AAG?TTG?GGT
GCT?CCA?TCT?ATT?ACT?TGT?GTT?AGA?AGA?GCT?TTC?GCT?TTG?GAA?TGT
ATT?AGA?GCT?ATT?GCT?GAG?AAG?AAG?GCT?GAT?GCT?GTT?ACT?TTG?GAT
GGT?GGT?ATG?GTT?TTC?GAA?GCT?GGT?AGA?GAT?CCA?TAC?AAG?TTG?AGA
CCA?GTT?GCT?GCT?GAA?ATT?TAC?GGT?ACT?AAG?GAA?TCT?CCA?CAA?ACT
CAT?TAC?TAC?GCT?GTT?GCT?GTT?GTT?AAG?AAG?GGT?TCT?AAC?TTT?CAA
TTG?GAT?CAA?TTG?CAA?GGT?AGA?AAG?TCT?TGT?CAT?ACT?GGT?TTG?GGT
AGA?TT
The aminoacid sequence of this Bovinelactoferrin peptide is:
aprknvrwct?isqpewfkcr?rwqwrmkklg?apsitcvrra?faleciraia?ekkadavtld?ggmvfeagrdpyklrpvaae?iygtkespqt?hyyavavvkk?gsnfqldqlq?grkschtglgr。
2. Bovinelactoferrin peptide according to claim 1 is characterized in that, the active centre of described Bovinelactoferrin peptide nucleotide sequence is:
TTT?AAG?TGT?AGA?AGA?TGG?CAA?TGG?AGA?ATG?AAG?AAG?TTG?GGT?GCT
CCA?TCT?ATT?ACT?TGT?GTT?AGA?AGA?GCT?TTC
The active centre of this Bovinelactoferrin peptide ammino acid sequence is:
fkcrrwqwrm?kklgapsitc?vrraf。
3. the preparation method of claim 1 or 2 described Bovinelactoferrin peptides is characterized in that, may further comprise the steps:
1) structure of recombinant expression vector;
2) abduction delivering fusion rotein;
3) isolation and purification of protein.
4. the preparation method of Bovinelactoferrin peptide according to claim 3 is characterized in that: described step 1) comprising:
Synthesizing of Bovinelactoferrin peptide gene;
Bovinelactoferrin peptide gene cloning;
Make up recombinant expression vector.
5. the preparation method of Bovinelactoferrin peptide according to claim 4 is characterized in that: described step 2) comprising:
Contain the acquisition of the engineering bacteria of Bovinelactoferrin peptide gene;
The abduction delivering fusion rotein.
6. the preparation method of Bovinelactoferrin peptide according to claim 5 is characterized in that: described step 3) comprising:
The separation of fusion rotein;
Concentrating of fusion rotein;
The purifying of fusion rotein;
The preservation of Bovinelactoferrin peptide.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011102721788A CN102993296A (en) | 2011-09-14 | 2011-09-14 | Bovine lactoferricin and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011102721788A CN102993296A (en) | 2011-09-14 | 2011-09-14 | Bovine lactoferricin and preparation method thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN102993296A true CN102993296A (en) | 2013-03-27 |
Family
ID=47922460
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2011102721788A Pending CN102993296A (en) | 2011-09-14 | 2011-09-14 | Bovine lactoferricin and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102993296A (en) |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104073511A (en) * | 2014-07-02 | 2014-10-01 | 广东希普生物科技股份有限公司 | Bovine lactoferricin peptide constitutive type yeast expression vector and preparation method thereof |
| CN104313048A (en) * | 2014-10-21 | 2015-01-28 | 长沙中科晶博生物科技有限公司 | Method for producing lactoferrin by using saccharomyces cerevisiae |
| CN105177097A (en) * | 2015-10-21 | 2015-12-23 | 哈尔滨工业大学 | Preparation method and application of lactoferricin for promoting proliferative activity of bone cells |
| CN105274137A (en) * | 2015-11-13 | 2016-01-27 | 长沙中科晶博生物科技有限公司 | Method of producing lactoferrin by using Artemisia apiacea |
| CN106377762A (en) * | 2016-08-24 | 2017-02-08 | 方雅悯 | Application of bovine lactoferrin and zymolytes thereof in products for protecting stomach and liver |
| CN106519022A (en) * | 2016-11-16 | 2017-03-22 | 成都中医药大学 | Recombinant bovine lactoferricin derived peptide, and preparation method and applications thereof |
| CN108929866A (en) * | 2018-06-11 | 2018-12-04 | 黑龙江八农垦大学 | The new function and its antibacterial peptide of bacillus subtilis GGT protein degradation product are identified |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101395180A (en) * | 2005-12-30 | 2009-03-25 | 赢创罗姆有限责任公司 | Peptides useful as cell-penetrating peptides |
| US20090202574A1 (en) * | 2004-11-19 | 2009-08-13 | Jagat Rakesh Kanwar | Methods of immune or haematological enhancement, inhibiting tumour formation or growth, and treating or preventing cancer |
| US20090270309A1 (en) * | 2005-10-14 | 2009-10-29 | Jillian Cornish | Use of lactoferrin fragments and hydrolysates |
| CN101628941A (en) * | 2008-07-18 | 2010-01-20 | 中国农业大学 | Bovine lactoferrin antibacterial peptide fusion protein, coding gene and application thereof |
| CN101701242A (en) * | 2009-11-26 | 2010-05-05 | 东北农业大学 | Method for separating bovine lactoferricin |
-
2011
- 2011-09-14 CN CN2011102721788A patent/CN102993296A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20090202574A1 (en) * | 2004-11-19 | 2009-08-13 | Jagat Rakesh Kanwar | Methods of immune or haematological enhancement, inhibiting tumour formation or growth, and treating or preventing cancer |
| US20090270309A1 (en) * | 2005-10-14 | 2009-10-29 | Jillian Cornish | Use of lactoferrin fragments and hydrolysates |
| CN101395180A (en) * | 2005-12-30 | 2009-03-25 | 赢创罗姆有限责任公司 | Peptides useful as cell-penetrating peptides |
| CN101628941A (en) * | 2008-07-18 | 2010-01-20 | 中国农业大学 | Bovine lactoferrin antibacterial peptide fusion protein, coding gene and application thereof |
| CN101701242A (en) * | 2009-11-26 | 2010-05-05 | 东北农业大学 | Method for separating bovine lactoferricin |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104073511A (en) * | 2014-07-02 | 2014-10-01 | 广东希普生物科技股份有限公司 | Bovine lactoferricin peptide constitutive type yeast expression vector and preparation method thereof |
| CN104313048A (en) * | 2014-10-21 | 2015-01-28 | 长沙中科晶博生物科技有限公司 | Method for producing lactoferrin by using saccharomyces cerevisiae |
| CN105177097A (en) * | 2015-10-21 | 2015-12-23 | 哈尔滨工业大学 | Preparation method and application of lactoferricin for promoting proliferative activity of bone cells |
| CN105177097B (en) * | 2015-10-21 | 2019-04-09 | 哈尔滨工业大学 | A kind of preparation method with the lactoferricin for promoting activity of osteoblast proliferation |
| CN105274137A (en) * | 2015-11-13 | 2016-01-27 | 长沙中科晶博生物科技有限公司 | Method of producing lactoferrin by using Artemisia apiacea |
| CN105274137B (en) * | 2015-11-13 | 2018-12-25 | 长沙中科晶博生物科技有限公司 | A method of lactoferrin is produced with sweet wormwood |
| CN106377762A (en) * | 2016-08-24 | 2017-02-08 | 方雅悯 | Application of bovine lactoferrin and zymolytes thereof in products for protecting stomach and liver |
| CN106519022A (en) * | 2016-11-16 | 2017-03-22 | 成都中医药大学 | Recombinant bovine lactoferricin derived peptide, and preparation method and applications thereof |
| CN108929866A (en) * | 2018-06-11 | 2018-12-04 | 黑龙江八农垦大学 | The new function and its antibacterial peptide of bacillus subtilis GGT protein degradation product are identified |
| CN108929866B (en) * | 2018-06-11 | 2021-04-06 | 黑龙江八一农垦大学 | New function of Bacillus subtilis GGT protein degradation product and identification of antibacterial peptide thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102993296A (en) | Bovine lactoferricin and preparation method thereof | |
| CN102127549B (en) | Chicken antimicrobial peptides Cathelicidins and preparation method and applications thereof | |
| CN105061603B (en) | Antibacterial peptide-lysozyme fusion protein preparation method and applications | |
| CN108794635B (en) | Bovine lactoferricin-human lysozyme fusion protein, gene and application thereof | |
| CN103554225B (en) | Synthetic antibacterial peptides and application thereof | |
| CN106148455B (en) | Inducer and application thereof | |
| CN101817883A (en) | Housefly cecropin-human lysozyme fusion protein, and preparation method and application thereof | |
| CN103789317A (en) | Scylla paramamosain antibacterial peptide hyastatin gene clone, encoding protein recombination and application | |
| CN114015698B (en) | Recombinant lactococcus lactis expressing heat shock protein Hsp70 and preparation method and application thereof | |
| CN107446941A (en) | Cecropin A antibacterial peptide based on self-aggregation short-peptide induction and preparation method thereof | |
| CN102586262B (en) | Defensin gene of antimicrobial peptide of bemisia tabaci (Gennadius), antimicrobial peptide encoded by defensin gene and preparation method for defensin gene | |
| CN104250304B (en) | The vaccine combination of a kind of fusion protein and its coding and application | |
| CN103103196A (en) | Modified goat defensin gene and preparation method and application thereof | |
| Peng et al. | Tissue distribution, expression, and antimicrobial activity of Anas platyrhynchos avian β-defensin 6 | |
| CN110256570A (en) | A kind of recombination fusion antibacterial peptide and application | |
| CN102240399B (en) | Application of siniperca chuatsi ISKNV (Infectious Spleen and Kidney Necrosis Virus) ORF093 protein | |
| CN101985467A (en) | Analgesic active peptide VGG and preparation method and applications thereof | |
| WO2011060613A1 (en) | Human alpha-defensin 5 mutant polypeptide and preparation method and use thereof | |
| CN104313048A (en) | Method for producing lactoferrin by using saccharomyces cerevisiae | |
| CN110478480A (en) | Sore mouth virus F1L vaccine based on ferritin nano particle and preparation method thereof | |
| CN101475942B (en) | B19 virus VP1 unique region gene | |
| CN102586257A (en) | Sika deer IGF (Insulin-like Growth Factor)-1 polypeptide and preparation method thereof | |
| CN105713908A (en) | Recombinant bombyx mori gloverin and preparation method and application thereof | |
| CN103830747A (en) | Genetically engineered vaccine of epsilon toxin of clostridium perfringens and application thereof | |
| CN104402974B (en) | A kind of polypeptide with mucosal adjuvant activity and its purposes in mucosal adjuvant is prepared |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20130327 |