CN104560865A - Cell line resistant to A subgroup avian leukosis virus as well as construction method and application thereof - Google Patents
Cell line resistant to A subgroup avian leukosis virus as well as construction method and application thereof Download PDFInfo
- Publication number
- CN104560865A CN104560865A CN201410831496.7A CN201410831496A CN104560865A CN 104560865 A CN104560865 A CN 104560865A CN 201410831496 A CN201410831496 A CN 201410831496A CN 104560865 A CN104560865 A CN 104560865A
- Authority
- CN
- China
- Prior art keywords
- alv
- subgroup avian
- env
- clone
- subgroup
- 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.)
- Granted
Links
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention discloses a cell line resistant to an A subgroup avian leukosis virus as well as a construction method and application thereof, and belongs to the technical field of agricultural biology. The cell line resistant to the A subgroup avian leukosis virus is named after an anti ALV-A chicken embryo fibroblast cell line DF-1/A, and is conserved in Wuhan University, China of China Center for Type Culture Collection (CCTCC) with the preservation number of CCTCC NO: C2014180 from October 15th, 2014; the cell line resistant to the A subgroup avian leukosis virus is applied to the diagnosis of the A subgroup avian leukosis virus. The cell line resistant to the A subgroup avian leukosis virus disclosed by the invention can achieve stable passage and be stored for a long term, is helpful for accurately identifying subgroup classification of ALV by using the characteristic of ALV superinfection, and is applied to the diagnosis of the A subgroup avian leukosis virus.
Description
Technical field
The invention belongs to agricultural biological technical field, particularly a kind of clone of anti-A subgroup avian leucosis virus and construction process thereof and application.
Background technology
Current avian leukosis is serious to the harm of China's aviculture, and has occurred the multiple pathological manifestations types such as lymphoma, myelocytome type and angiomatous type, and clinical manifestation is also easily obscured with the neoplastic disease such as reticuloendotheliosis disease, Marek mutually.This disease is mainly vertical transmission, control these sick effective measures be reduce breeder flock infection rate and set up without leukemic breeder flock, its Main Means is that the frequency by regularly carrying out exogenous ALV to chicken group detects, and eliminates the malicious positive chicken of band in time, to purify and to eliminate the ALV in chicken group.In exogenous ALV, have significantly pathogenic to chicken with J subgroup, A subgroup and B subgroup avian leucosis virus, also large to the harm of avian production.
At present, the domestic and international detection for avian leukosis cause of disease and authentication method mainly contain cell cultures, ELISA, indirect immunofluorescence assay, PCR etc., wherein ELISA is most widely used clinically, its advantage is this method operation relative ease, have commercial detection kit to utilize, its weak point to distinguish endogenous and exogenous ALV; Indirect immunofluorescence assay is relatively more directly perceived, but often needs to use specific antibody (monoclonal antibody).PCR primer and the work system thereof that can distinguish different ALV subgroup have report.
In the separation of exogenous avian leukosis virus, cell conventional at present has chick embryo fibroblast (CEF) and comes from the fibroblast DF-1 that 0 is chicken.U.S. ADOL laboratory, based on DF-1 clone, establishes the DF-1/J clone of J substock lymphoid leuoosis-resistant poison.Although there are some specific chicken strains in U.S. ADOL laboratory, its primary cell produced can limit specific exogenous avian leukosis virus growth respectively, the rarity performance limiting its usefulness of these Strains of Chickens.At present, the relevant report that can limit the clone of A subgroup avian leucosis virus specifically is not also had.
Summary of the invention
For overcoming the shortcoming of prior art with not enough, primary and foremost purpose of the present invention is the clone providing a kind of anti-A subgroup avian leucosis virus.
Another object of the present invention is to the construction process of the clone that above-mentioned anti-A subgroup avian leucosis virus is provided.
Another object of the present invention is to provide the application of the clone of described anti-A subgroup avian leucosis virus.
Object of the present invention is achieved through the following technical solutions: a kind of clone of anti-A subgroup avian leucosis virus, the anti-ALV-A chick embryo fibroblast system DF-1/A of called after; Be preserved in China typical culture collection center; Preservation address is Wuhan, China Wuhan University; Deposit number is CCTCC NO:C2014180; Preservation from date is on October 15th, 2014.
The construction process of the clone of above-mentioned anti-A subgroup avian leucosis virus, comprises the steps:
(1) primer of band BamHI and NotI restriction enzyme site is utilized to increase env fragment from the plasmid RCAS (A) containing ALV-A env gene, respectively double digestion is carried out to the env gene increased and eukaryon expression plasmid pcDNA3.1/Zeo (+) (purchased from life company) with restriction enzyme BamHI and NotI, after purifying, A subgroup avian leucosis virus (ALV-A env) gene fragment is connected to pcDNA3.1/Zeo (+) eukaryon expression plasmid respectively, is built into recombinant plasmid pcDNA-env-A;
(2) on the basis that transfer vector pcDNA-env-A expresses in DF-1 chick embryo fibroblast system, repeatedly screened by Zeocin medicine, obtain the chick embryo fibroblast system DF-1/A that there is Zeocin resistance, Absorbable organic halogens expresses ALV-A env albumen, i.e. the clone of anti-A subgroup avian leucosis virus.
The primer of the band BamHI described in step (1) and NotI restriction enzyme site is respectively:
Upstream primer F:CGCGGATCCGCCACCATGGAAGCCGTCATTAAGGCATTTCTGACTGGATACCCT;
Downstream primer R:AAGGAAAAAAGCGGCCGCTTATACTATTCTGCTT.
The plasmid RCAS (A) containing ALV-A env gene described in step (1) derives from U.S. ADOL laboratory, specifically see such as Publication about Document: Hughes S H.The RCAS vector system [J] .Folia Biol (Praha) .2004,50 (3-4): 107-119.
The treatment process that Zeocin medicine described in step (2) repeatedly screens was specially with 200 μ g/mL zeocin medicines cell step sizing 30 generation.
The clone of described anti-A subgroup avian leucosis virus can be applicable to the diagnosis of A subgroup avian leucosis virus.
Invention mechanism of the present invention: avian leukosis virus envelope protein (the viral glycosylated protein of env genes encoding) is the major surface protein of this viroid, it determines the host range of virus infection, can induce and produce neutralizing antibody.This albumen and the identification of target cells specific receptors, combination, be cell entry cell, copy necessary.After infecting specific subgroup ALV, the viral glycosylated protein of the env genes encoding that cell surface virus receptor can be produced by this virus is closed, and shows powerful superingection resistance, can limit the subinfection again of corresponding virus.The present invention is first from env albumen superingection resistance, A subgroup avian leucosis virus env gene is inserted pcDNA3.1/Zeo (+) construction of eukaryotic expression vector and becomes transfer vector pcDNA-env-A, then by carrier pcDNA-env-A transfection chick embryo fibroblast system DF-1, screened by medicine Zeocin, obtain the clone of anti-A subgroup avian leucosis virus of anti-Zeocin, stably express ALV-A env albumen; Detected by the env genetic expression of method to DF-1/A clone such as PCR, indirect immunofluorescence (IFA) and Western-blot, and carried out anti-virus infection detection.
The present invention has following advantage and effect relative to prior art:
The present invention is from env albumen superingection resistance, A subgroup avian leucosis virus env gene is inserted pcDNA3.1/Zeo (+) construction of eukaryotic expression vector and becomes transfer vector pcDNA-env-A, then by carrier pcDNA-env-A transfection chick embryo fibroblast system DF-1, screened by medicine Zeocin, obtain the clone of anti-A subgroup avian leucosis virus of anti-Zeocin, stably express ALV-A env albumen.The clone of the anti-A subgroup avian leucosis virus obtained can be stablized and goes down to posterity, and preserves for a long time; Utilize the characteristic of ALV superingection, the subgroup classification accurately differentiating ALV can be contributed to, can be applicable to the diagnosis of A subgroup avian leucosis virus.
Accompanying drawing explanation
Fig. 1 is the cytological map of anti-A subgroup avian leucosis virus.
Fig. 2 is that PCR detects env genetic results figure in DF-1/A, wherein: M is Marker, DF-1 is negative control group, and DF-1/A is positive group.
Fig. 3 is the fluorescence results figure that indirect immunofluorescence (IFA) detects env expression conditions, and wherein: A is DF-1 groups of cells, B is DF-1/A groups of cells.
Fig. 4 is the result figure that Western-blot detects the expression of env gene.
Fig. 5 is the Antiviral breeding detected result figure of anti-ALV-A RAV-1 and ALV-A GD13 strain, and wherein: A is anti-ALV-A RAV-1 strain, B is anti-ALV-A GD13 strain.
Fig. 6 is the Antiviral breeding detected result figure of anti-ALV-J CHN06 strain
Fig. 7 is that clinical virus is hived off diagnostic result figure.
Embodiment
Below in conjunction with embodiment and accompanying drawing, the present invention is described in further detail, but embodiments of the present invention are not limited thereto.
Present invention employs following technical scheme:
DF-1 clone: from live vaccine initiative key lab of the Ministry of Agriculture.
Strain: RAV-1 and GD13 is ALV-A, CHN06 is ALV-J, and live vaccine initiative key lab of the Ministry of Agriculture provides.
Enzyme: restriction enzyme BamHI, NotI and T4 ligase enzyme is purchased from NEB company; LA Taq is purchased from TaKaRa company.
Plasmid: eukaryon expression plasmid pcDNA3.1/Zeo (+) is purchased from Invitrogen company; RCAS (A) plasmid, U.S. ADOL laboratory is presented.
Antibody: ALV-A monoclonal antibody specific, U.S. ADOL laboratory is presented.The goat anti-mouse IgG antibodies of FITC mark and the goat anti-mouse IgG antibodies of HRP mark are all purchased from Simga company.
Test kit: SQ Tissue tissue DNA extracts test kit, glue reclaims test kit DNA Gel Extraction Kit, goes intracellular toxin plasmid extraction test kit Plasmid Miniprep Kit to be OMEGA Products.ALV antigen detection kit (Avian Leukosis Virus Antigen Test Kit) is American I DEXX Products.
Cell culture: DMEM in high glucose medium powder, 0.25% pancreatin (containing EDTA), foetal calf serum are U.S. GIBCO Products; Dimethyl sulfoxide (DMSO) (DMSO) is Sigma Products.
The present invention increases the upstream and downstream primer (F and R) of ALV-A env gene by contriver's designed, designed:
F:CGCGGATCCGCCACCATGGAAGCCGTCATTAAGGCATTTCTGACTGGATACCCT
R:AAGGAAAAAAGCGGCCGCTTATACTATTCTGCTT
Reaction system (50 μ L): each 1 μ L, the ddH of LA Taq 25 μ L, upstream and downstream primer
2o 22.5 μ L, template 0.5 μ L.
Response procedures: 95 DEG C of denaturation 5min; 95 DEG C of sex change 30s, 57 DEG C of annealing 30s, 72 DEG C extend 2min, 30 circulations; 8min is extended after 72 DEG C.
From RCAS (A) plasmid, amplify env fragment with the primer of designed, designed, reclaim test kit DNA Gel Extraction Kit specification sheets according to glue and reclaim; Respectively double digestion is carried out to the env fragment of eukaryon expression plasmid pcDNA3.1/Zeo (+) and recovery according to NEB company restriction enzyme BamHI, NotI specification sheets; Reclaim digestion products respectively, and according to NEB company's T 4 ligase enzyme specification sheets, recovery product is connected, construction recombination plasmid pcDNA-env-A; Transformed competence colibacillus cell DH5 α, removes intracellular toxin plasmid extraction test kit specification sheets according to OMEGA company, carries out recombinant plasmid pcDNA-env-A extracting.Plasmid send life company to check order.
Check order correct plasmid according to Invitrogen
tMcompany
2000 transfection reagent box specification sheetss carry out transfection.
The DF-1 cell of digestion early-stage preparations, spreads six porocyte culture plates, 10%FBS, 5%CO
2, 37 DEG C cultivate.Carry out transfection in 24 hours, Corning company six porocyte culture plate every hole rotaring redyeing system is: 4 μ gpcDNA-env-A plasmids and 10 μ L liposomes use 250 μ L respectively
dilute and hatch 5min, and then mixing, jointly hatching 20min, 500 μ L plasmid-liposome complex being joined in ready six porocytes cultivations.After transfection, 5h changes cell culture fluid (DMEM+10%FBS), after 48h, the cell dissociation in six orifice plates hole is got off, spread 24 orifice plates, every hole 500 μ L cell culture fluid (DMEM+15%FBS+200 μ g/mL zeocin), changed once the cell culture fluid of same system, within 10-15 days, occurs single cell colonies every 3-4 days, about three weeks, single cell colonies is grown up, and digests to be placed in 6 orifice plates to cultivate, adherent cover with after proceed to 25cm
2cultivate in cell bottle, now cell culture is (DMEM+10%FBS+200 μ g/mL zeocin).With medicine zeocin to cell step sizing 30 generation, afterwards with the culture medium culturing cell not containing zeocin, from nucleic acid and protein level, intracellular env is detected, and carry out the application of Antiviral breeding and Virus Type.
Cell state: threadiness, fusiformis, adherent growth.(as shown in Figure 1)
PCR detects env gene in DF-1/A: the DNA of extracting DF-1/A, and according to the primer of aforementioned amplification env gene, system and response procedures amplification env gene, as shown in Figure 2, env gene amplification fragment size is 1844bp to result.
Indirect immunofluorescence (IFA) detects env expression conditions: in 24 orifice plates of confluent monolayers DF-1/A cell, and after cultivating 5d, bottom cell PBS washes 3 times, fixes 20min with 4% paraformaldehyde.Wash 3 times with PBS again, add the ALV-A monoclonal antibody of 1:200 dilution, 4 DEG C of overnight incubation.PBS washs 3 times, adds the sheep anti-mouse igg-FITC fluorescence antibody that 1:200 dilutes, and after 37 DEG C of effects 1h, PBS wash 3 times, adds 1 drop volume mark 50% glycerine and covers cell, observation experiment result under fluorescent microscope.Set up a DMEM negative control.As shown in Figure 3, there is specific fluorescence (Fig. 3 B) under fluorescent microscope, can be observed DF-1/A cell endochylema and surface to result, and DF-1 cell does not have green fluorescence (Fig. 3 A).External source env fragment successful expression in DF-1/A is described.
Western-blot detects the expression of env gene: carry total protein of cell after DF-1/A and contrast DF-1 cultivates 5d, electrophoresis is carried out with Bio-Rad vertical electrophoresis apparatus, concentrated glue voltage is 80V, separation gel electrophoretic voltage is 100V, electrophoretic buffer is 1 × Tris-glycine running buffer, reaches after bottom separation gel stop electrophoresis until bromophenol blue indicator.After cutting glue transferring film, NC film 1 × PBS is washed 3 times, each 5min.NC film is placed in the PBS of skim-milk containing 5%, room temperature closes 2h, and 1 × PBS washs 3 times, each 5min.Add ALV-A monoclonal antibody and Actin monoclonal antibody, hatch 12h for 4 DEG C.Antibody dilution concentration is ALV-A monoclonal antibody (1:500), Actin (1:1000).After primary antibodie is hatched, film PBST is washed 3 times, each 5min, the IgG bis-NC film being placed in 1:100000HRP mark resists, and hatch 1h for 37 DEG C, 1 × PBST washs 3 times, each 5min.In darkroom, add ECL immunochemiluminescence liquid and hatch 5min, wrap NC film be placed on sensitive film with preservative film on NC film, development, fixing, picture scanning after drying, preserves.Result as shown in Figure 4, has env protein expression, and does not have in DF-1 in DF-1/A.External source env fragment successful expression in DF-1/A is described.
Antiviral breeding: respectively with DMEM by ALV-A strain RAV-1 and GD13 from 10
0to 10
5carry out doubling dilution.Digest DF-1/A cell and DF-1 cell respectively, be respectively inoculated in one piece of 24 porocyte culture plate, every hole inoculation 1mL cell suspension, cell concn is 1.7 × 10
5cells/mL.Be inoculated in DF-1/A and DF-1 cell suspension respectively by the venom of each for RAV-1 and GD13 viral dilution gradient, 100 μ L venom are inoculated in every hole, and each viral dilution gradient does three repetitions, and do contrast and the DMEM negative control of a known positive strain, 5%CO
2, 10%FBS, to cultivate in 37 DEG C of incubators.After venom to be seeded second day, after cell covers with individual layer, the cell maintenance medium being changed to 1%FBS was cultivated.Cultured continuously 6d, period continues to monitor the growing state of cell, and at the 6th day collecting cell culture, centrifugal collecting cell supernatant after freeze thawing three times.The cell conditioned medium liquid the collected ELISA kit of same batch carries out the detection of p27 antigen ELISA.As shown in Figure 5, RAV-1 and GD13 normally can breed result on DF-1, but on DF-1/A, all can not normally breed, and is only 10 at virus titer
5tCID
50copying appears in time, but virus replication is obviously suppressed, and is starkly lower than copying on DF-1.Result illustrates, DF-1/A has the ability of very strong anti-ALV-A infected cell.Use the same method ALV-J strain CHN06 inoculation DF-1/A cell and DF-1 cell.Result such as Fig. 6 shows, and ALV-J can normally breed on DF-1/A and DF-1, and breeding situation does not have difference, illustrates that DF-1/A does not have resistance to ALV-J.
Clinical virus is hived off diagnosis example: as shown in Figure 7, by ALV Simultaneous vaccination DF-1 and DF-1/A of clinical separation, cultivate after 7 days, centrifugal collecting cell supernatant after freeze thawing three times, ELISA detects, the s/p value of DF-1/A is starkly lower than DF-1, illustrates that DF-1/A has resistance to this ALV, tentatively judges that this ALV is ALV-A.The DF-1 cell DNA of this ALV is inoculated in extracting, and amplification gp85, send order-checking after recovery, prove that this strain is really ALV-A.
Above-described embodiment is the present invention's preferably embodiment; but embodiments of the present invention are not restricted to the described embodiments; change, the modification done under other any does not deviate from spirit of the present invention and principle, substitute, combine, simplify; all should be the substitute mode of equivalence, be included within protection scope of the present invention.
Claims (5)
1. a clone for anti-A subgroup avian leucosis virus, is characterized in that, the anti-ALV-A chick embryo fibroblast system DF-1/A of described clone called after; Be preserved in China typical culture collection center; Preservation address is Wuhan, China Wuhan University; Deposit number is CCTCC NO:C2014180; Preservation from date is on October 15th, 2014.
2. the construction process of the clone of anti-A subgroup avian leucosis virus according to claim 1, is characterized in that: comprise the steps:
(1) primer of band BamHI and NotI restriction enzyme site is utilized to increase env fragment from the plasmid RCAS (A) containing ALV-A env gene, respectively double digestion is carried out to the env gene increased and eukaryon expression plasmid pcDNA3.1/Zeo (+) with restriction enzyme BamHI and NotI, after purifying, ALV-A env gene fragment is connected to pcDNA3.1/Zeo (+) eukaryon expression plasmid respectively, is built into recombinant plasmid pcDNA-env-A;
(2) on the basis that transfer vector pcDNA-env-A expresses in DF-1 chick embryo fibroblast system, repeatedly screened by Zeocin medicine, obtain the chick embryo fibroblast system DF-1/A that there is Zeocin resistance, Absorbable organic halogens expresses ALV-A env albumen, i.e. the clone of anti-A subgroup avian leucosis virus.
3. the construction process of the clone of anti-A subgroup avian leucosis virus according to claim 2, is characterized in that: the primer of the band BamHI described in step (1) and NotI restriction enzyme site is respectively:
Upstream primer F:CGCGGATCCGCCACCATGGAAGCCGTCATTAAGGCATTTC TGACTGGATACCCT;
Downstream primer R:AAGGAAAAAAGCGGCCGCTTATACTATTCTGCTT.
4. the construction process of the clone of anti-A subgroup avian leucosis virus according to claim 2, is characterized in that: the treatment process that the Zeocin medicine described in step (2) repeatedly screens was specially with 200 μ g/mL zeocin medicines cell step sizing 30 generation.
5. the clone of anti-A subgroup avian leucosis virus according to claim 1 is applied to the diagnosis of A subgroup avian leucosis virus.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410831496.7A CN104560865B (en) | 2014-12-25 | 2014-12-25 | The cell line of anti-A subgroup avian leucosis virus and its construction method and application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410831496.7A CN104560865B (en) | 2014-12-25 | 2014-12-25 | The cell line of anti-A subgroup avian leucosis virus and its construction method and application |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104560865A true CN104560865A (en) | 2015-04-29 |
| CN104560865B CN104560865B (en) | 2018-05-01 |
Family
ID=53077945
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410831496.7A Active CN104560865B (en) | 2014-12-25 | 2014-12-25 | The cell line of anti-A subgroup avian leucosis virus and its construction method and application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104560865B (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105861444A (en) * | 2016-04-08 | 2016-08-17 | 华南农业大学 | Anti-subgroup B avian leukosis virus strain cell line and application thereof |
| CN105861445A (en) * | 2016-04-08 | 2016-08-17 | 华南农业大学 | Cell system for preventing K avian leukosis virus (ALV-K) subgroup and application of cell system |
| CN108896765A (en) * | 2018-07-04 | 2018-11-27 | 山东农业大学 | A kind of colloidal gold strip and preparation method thereof detecting A subgroup avian leucosis virus |
| CN110358733A (en) * | 2019-05-24 | 2019-10-22 | 中国农业科学院哈尔滨兽医研究所（中国动物卫生与流行病学中心哈尔滨分中心） | One plant is stablized the cell line and application for expressing A subgroup avian leucosis virus gp85 albumen |
| CN114032216A (en) * | 2021-10-22 | 2022-02-11 | 山东农业大学 | New application of bisporticosteroid-like kinase 1 |
-
2014
- 2014-12-25 CN CN201410831496.7A patent/CN104560865B/en active Active
Non-Patent Citations (3)
| Title |
|---|
| CHESTERS P.M,ET AL: "EM:AY608692.1;Avian leukosis virus envelope(env)gene,complete cds", 《EBI》 * |
| 叶建强等: "J亚群禽白血病度超感染抗性鸡胚成纤维细胞系建立", 《中国畜牧兽医学会家畜传染病学分会第六届全国会员代表大会暨第11次学术讨论会论文集》 * |
| 张小桃等: "A亚群禽白血病病毒GD08株的分离与全基因组序列测定", 《中国兽医学报》 * |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105861444A (en) * | 2016-04-08 | 2016-08-17 | 华南农业大学 | Anti-subgroup B avian leukosis virus strain cell line and application thereof |
| CN105861445A (en) * | 2016-04-08 | 2016-08-17 | 华南农业大学 | Cell system for preventing K avian leukosis virus (ALV-K) subgroup and application of cell system |
| CN105861444B (en) * | 2016-04-08 | 2019-08-06 | 华南农业大学 | The cell line and its application of one plant of anti-B subgroup avian leucosis virus |
| CN105861445B (en) * | 2016-04-08 | 2019-08-06 | 华南农业大学 | The cell line and its application of one plant of anti-K subgroup avian leucosis virus |
| CN108896765A (en) * | 2018-07-04 | 2018-11-27 | 山东农业大学 | A kind of colloidal gold strip and preparation method thereof detecting A subgroup avian leucosis virus |
| CN110358733A (en) * | 2019-05-24 | 2019-10-22 | 中国农业科学院哈尔滨兽医研究所（中国动物卫生与流行病学中心哈尔滨分中心） | One plant is stablized the cell line and application for expressing A subgroup avian leucosis virus gp85 albumen |
| CN110358733B (en) * | 2019-05-24 | 2022-02-15 | 中国农业科学院哈尔滨兽医研究所（中国动物卫生与流行病学中心哈尔滨分中心） | Cell line for stably expressing subgroup A avian leukosis virus gp85 protein and application |
| CN114032216A (en) * | 2021-10-22 | 2022-02-11 | 山东农业大学 | New application of bisporticosteroid-like kinase 1 |
| CN114032216B (en) * | 2021-10-22 | 2023-07-14 | 山东农业大学 | New use of double-cortex epinephrine-like kinase 1 |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104560865B (en) | 2018-05-01 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Johne et al. | Characterization of two novel polyomaviruses of birds by using multiply primed rolling-circle amplification of their genomes | |
| Pedrera et al. | Bovine herpesvirus-4-vectored delivery of Nipah virus glycoproteins enhances T cell immunogenicity in pigs | |
| Usherwood et al. | Latent antigen vaccination in a model gammaherpesvirus infection | |
| CN104560865A (en) | Cell line resistant to A subgroup avian leukosis virus as well as construction method and application thereof | |
| He et al. | The use of an in vitro microneutralization assay to evaluate the potential of recombinant VP5 protein as an antigen for vaccinating against Grass carp reovirus | |
| Noh et al. | Isolation and characterization of novel bat paramyxovirus B16-40 potentially belonging to the proposed genus Shaanvirus | |
| CN104531700B (en) | Suppress shRNA sequences and its application of mouse MACF1 gene expressions | |
| CN111450244A (en) | Cell composition for preventing and treating coronavirus infection and application thereof | |
| CN105861445B (en) | The cell line and its application of one plant of anti-K subgroup avian leucosis virus | |
| Wei et al. | Strain-dependent porcine circovirus type 2 (PCV2) entry and replication in T-lymphoblasts | |
| Li et al. | Characterization of Ictalurid herpesvirus 1 glycoprotein ORF59 and its potential role on virus entry into the host cells | |
| Jiao et al. | Construction and immunogenicity of virus-like particles of feline parvovirus from the tiger | |
| CN102732486B (en) | Porcine circovirus type2 strain and application thereof | |
| CN106018780A (en) | F1-protein-based indirect immunofluorescence kit for detecting type 4 fowl adenovirus antibody | |
| Liu et al. | Recombinant bovine herpesvirus type I expressing the bovine viral diarrhea virus E2 protein could effectively prevent infection by two viruses | |
| CN110195080A (en) | Stablize recombinant plasmid, recombinant slow virus and porcine alveolar macrophage system and its construction method of expression CD163 receptor protein | |
| Shah et al. | Construction of a Novel Infectious Clone of Recombinant Herpesvirus of Turkey Fc-126 Expressing VP2 of IBDV | |
| CN105713866A (en) | Human cytomegalovirus (HCMV) infectious clone as well as construction method and applications of HCMV infectious clone | |
| CN103333865B (en) | HPV pseudovirus, kit thereof and method for detecting HPV neutralizing antibodies | |
| McNally et al. | Endogenous expression of ASLV viral proteins in specific pathogen free chicken embryos: relevance for the developmental biology research field | |
| CN108359645A (en) | A kind of preparation and application of expression II type capsid protein baculoviral of rabbit hemorrhagic disease virus | |
| Zeng et al. | Characterization of a recombinant thermostable Newcastle disease virus (NDV) expressing glycoprotein gB of infectious laryngotracheitis virus (ILTV) protects chickens against ILTV challenge | |
| CN103333916A (en) | Construction method and application of BHK cell line suitable for newcastle disease virus proliferation | |
| CN104232587A (en) | Cell line for stably expressing schmallenberg virus nucleocapsid protein and application of cell line | |
| Śmietanka et al. | A recombinant Turkey herpesvirus expressing F and HN genes of Avian Avulavirus-1 (AAvV-1) genotype VI confers cross-protection against challenge with virulent AAvV-1 genotypes IV and VII in chickens |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |