CN108715858A - A kind of nucleic acid molecules, protein and application thereof - Google Patents
A kind of nucleic acid molecules, protein and application thereof Download PDFInfo
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Abstract
The invention discloses a kind of nucleic acid molecules, protein and application thereof, can inhibit duplication of the virus in fish body, and its overexpression in fish body can significantly improve survival rate of the fish body during SVCV infects.
Description
Technical field
The present invention relates to biotechnologies more particularly to a kind of nucleic acid molecules, protein and application thereof.
Background technology
In high animal and plant cells, nadph oxidase is the main source for generating ROS, is mainly made of 5 subunits：
Gp91phox, p22phox, p40phox, p47phox, p67phox subunit；Wherein gp91phox is crucial subunit, shares 7 kinds
Hypotype, respectively by 7 genes, i.e. Nox family members nox1, nox2, nox3, nox4, nox5, Duox1 and Duox2 genes are compiled
Code.
The popularity of Nox family genes distribution determines the diversity of the family gene protein function, and research at present has been sent out
Existing nox albumen may participate in numerous physiology courses such as the growth, differentiation, the regulation and control of apoptosis and intercellular signal access of cell.
The effect about ROS in immune response has become the hot spot of research in recent years, is primarily involved in modulate host
The important physiology course such as defence capability and intracellular signal transduction；In the innate immunity, body oxidative burst generates a large amount of
ROS is the first line of defence for resisting cause of disease.Nox1 is the homologue of catalytic subunit gp91phox.At present studies have found that nox1
Mediate the ROS generated that can participate in the activation levels of regulation and control JNK；It may also participate in the receptor-mediated signals such as regulation and control TLR, NLRP3
Access.Under quiescent condition, the expression quantity of nox1 is relatively low；It can make Nox1 great expressions when there is the stimulations such as extraneous factor such as LPS
And activating transcription factor, and then the generation for promoting inflammatory factor and growth factor etc..
Huichun viremia virus (Spring Viraemia ofCarp Virus, SVCV) is to cause spring viremia
The cause of disease of (Spring Viraemia ofCarp, SVC).SVCV virus main infection objects are the carps such as grass carp, bighead and goldfish
Flying fish；Fish body local hemorrhage, slow in action, slow in reacting and long-time can be caused to stop or be gathered in after virus infection fish body
Basin bottom is until dead；Lethality is high and propagation is exceedingly fast, and often results in huge economic loss.Therefore, to can effectively control
The novel immune antiviral drugs of SVCV or the demand of recombinant vaccine are very urgent.
In bony fish, the only a small amount of report of the research in relation to nox1, and its study limitation nox1 expression with into
Change analysis.Although in mammals, existing research discloses inflammatory reaction and oxidation that nox1 can inhibit influenza virus to induce
Stress, but nox1 whether be conducive to host resist virus infect it is not clear.
Invention content
The defects of for the above-mentioned prior art, the present invention provides a kind of nucleic acid molecules, protein and application thereof, energy
Enough duplications for inhibiting virus in fish body, and its overexpression in fish body can significantly improve fish body depositing during SVCV infects
Motility rate.
The technical solution that the present invention is provided to solve above-mentioned technical problem is as follows：
On the one hand, a kind of nucleic acid molecules are provided comprising：(a) there is SEQ ID NO:Nucleotide sequence shown in 1；
Or
(b) there is SEQ ID NO:Nucleotide sequence shown in 1 can variant sequence thereof, and it is described can variant sequence thereof and SEQ ID
NO:Nucleotide sequence shown in 1 has at least 80% homology；Or
It (c) under strict conditions can be with the nucleotide sequence of the nucleotide sequence hybridization limited in (a)；Or
(d) there is SEQ ID NO:Nucleotide sequence shown in 2；Or
(e) there is SEQ ID NO:Nucleotide sequence shown in 2 can variant sequence thereof, and it is described can variant sequence thereof and SEQ ID
NO:Nucleotide sequence shown in 2 has at least 80% homology；Or
It (f) under strict conditions can be with the nucleotide sequence of the nucleotide sequence hybridization limited in (d)；Or
(g) coding has SEQ ID NO:The nucleotide sequence of the polypeptide of amino acid sequence shown in 3；Or
(h) nucleotide sequence of encoding amino acid sequence, and the amino acid sequence is SEQ ID NO:Amino acid shown in 2
The polypeptide sequence that sequence is formed after replacing, missing or adding one or more amino acid；Or
(i) coding has SEQ ID NO:The nucleotide sequence of the polypeptide of amino acid sequence shown in 4；Or
(j) nucleotide sequence of encoding amino acid sequence, and the amino acid sequence is SEQ ID NO:Amino acid shown in 4
The polypeptide sequence that sequence is formed after replacing, missing or adding one or more amino acid.
On the one hand, a kind of protein of separation is provided comprising：(a) there is SEQ ID NO:Amino acid sequence shown in 3
Protein；Or
(b) there is SEQ ID NO:Amino acid sequence shown in 3 is after replacing, missing or adding one or more amino acid
The protein of the sequence of formation；Or
(c) by with SEQ ID NO:The protein that the nucleic acid molecules expression of nucleotide sequence shown in 1 generates；Or
(d) protein generated by the nucleic acid molecules expression with nucleotide sequence, and the nucleotide sequence has
Under strict conditions with SEQ ID NO:The complementary series of nucleotide sequence hybridization shown in 1.
(e) there is SEQ ID NO:The protein of amino acid sequence shown in 4；Or
(f) there is SEQ ID NO:Amino acid sequence shown in 4 is after replacing, missing or adding one or more amino acid
The protein of the sequence of formation；Or
(g) by with SEQ ID NO:The protein that the nucleic acid molecules expression of nucleotide sequence shown in 2 generates；Or
(h) protein generated by the nucleic acid molecules expression with nucleotide sequence, and the nucleotide sequence has
Under strict conditions with SEQ ID NO:The complementary series of nucleotide sequence hybridization shown in 2.
On the one hand, a kind of recombinant protein is provided comprising destination protein and the polypeptide that can be inserted into the destination protein
Carrier, and the albumen that the destination protein is above-mentioned separation.
Preferably, the peptide carrier is prokaryotic expression carrier or carrier for expression of eukaryon.
On the one hand, a kind of polynucleotides are provided, are used to encode above-mentioned recombinant protein.
On the one hand, a kind of carrier is provided comprising above-mentioned nucleic acid molecules or polynucleotides.
Preferably, the carrier includes plasmid.
On the one hand, a kind of host cell is provided comprising above-mentioned nucleic acid molecules and/or above-mentioned carrier.
Preferably, the host cell includes fish cell.
On the one hand, a kind of pharmaceutical composition is provided comprising above-mentioned recombinant protein.
On the one hand, the purposes of aforementioned pharmaceutical compositions in medicine preparation is provided, the drug is for inhibiting viral sense
Dye.
Preferably, the virus includes huichun viremia virus.
Effect caused by technical solution of the present invention：
Nucleic acid molecules, the protein and application thereof of the present invention, can inhibit duplication of the virus in fish body, and it is in fish
Internal overexpression can significantly improve survival rate of the fish body during SVCV infects.
Description of the drawings
Fig. 1 is the electrophoretogram that nox1_tv1 is expanded in experimental example five of the present invention；
Fig. 2 be experimental example five of the present invention in nox1_tv1 recombinant plasmids fish body cell inner expression result figure；
Fig. 3 is the qRT-PCR result figures that nox1_tv1 is expressed in the fish body infected by SVCV in experimental example six of the present invention；
Fig. 4 is the result figure that nox1_tv1 influences SVCV-N in experimental example six of the present invention；
Fig. 5 is the result figure that nox1_tv1 influences SVCV-G in experimental example six of the present invention；
The result figure of juvenile fish Survival Effects after Fig. 6 infects virus for nox1_tv1 in experimental example six of the present invention；
Fig. 7 is the result figure that nox1_tv1 influences antiviral gene IFN1 in fish body in experimental example six of the present invention；
Fig. 8 is the result figure that nox1_tv1 influences antiviral gene MXA in fish body in experimental example six of the present invention.
Specific implementation mode
In order to make the purpose , technical scheme and advantage of the present invention be clearer, with reference to the accompanying drawings and embodiments, right
The present invention is further elaborated.It should be appreciated that the specific embodiments described herein are merely illustrative of the present invention, and
It is not used in the restriction present invention.
Embodiment one：
Present embodiments provide a kind of nucleic acid molecules comprising：(a) there is SEQ ID NO:Nucleotide sequence shown in 1；
Or (b) there is SEQ ID NO:Nucleotide sequence shown in 1 can variant sequence thereof, and it is described can variant sequence thereof and SEQ ID NO:1
Shown nucleotide sequence has at least 80% (preferably 80% or 82% or 85% or 88% or 90% or 92% or 93% or 95%
Or 97% or homology 99%)；Or
It (c) under strict conditions can be with the nucleotide sequence of the nucleotide sequence hybridization limited in (a)；Or
(d) there is SEQ ID NO:Nucleotide sequence shown in 2；Or
(e) there is SEQ ID NO:Nucleotide sequence shown in 2 can variant sequence thereof, and it is described can variant sequence thereof and SEQ ID
NO:Nucleotide sequence shown in 2 have at least 80% (preferably 80% or 82% or 85% or 88% or 90% or 92% or 93% or
95% or 97% or homology 99%)；Or
It (f) under strict conditions can be with the nucleotide sequence of the nucleotide sequence hybridization limited in (d)；Or
(g) coding has SEQ ID NO:The nucleotide sequence of the polypeptide of amino acid sequence shown in 3；Or
(h) nucleotide sequence of encoding amino acid sequence, and the amino acid sequence is SEQ ID NO:Amino acid shown in 2
The polypeptide sequence that sequence is formed after replacing, missing or adding one or more amino acid；Or
(i) coding has SEQ ID NO:The nucleotide sequence of the polypeptide of amino acid sequence shown in 4；Or
(j) nucleotide sequence of encoding amino acid sequence, and the amino acid sequence is SEQ ID NO:Amino acid shown in 4
The polypeptide sequence that sequence is formed after replacing, missing or adding one or more amino acid.
Embodiment two：
The present embodiment provides a kind of protein of separation comprising：(a) there is SEQ ID NO:Amino acid sequence shown in 2
Protein；Or
(b) there is SEQ ID NO:Amino acid sequence shown in 2 is after replacing, missing or adding one or more amino acid
The protein of the sequence of formation；Or
(c) by with SEQ ID NO:The protein that the nucleic acid molecules expression of nucleotide sequence shown in 1 generates；Or
(d) protein generated by the nucleic acid molecules expression with nucleotide sequence, and the nucleotide sequence has
Under strict conditions with SEQ ID NO:The complementary series of nucleotide sequence hybridization shown in 1；Or
(e) there is SEQ ID NO:The protein of amino acid sequence shown in 4；Or
(f) there is SEQ ID NO:Amino acid sequence shown in 4 is after replacing, missing or adding one or more amino acid
The protein of the sequence of formation；Or
(g) by with SEQ ID NO:The protein that the nucleic acid molecules expression of nucleotide sequence shown in 2 generates；Or
(h) protein generated by the nucleic acid molecules expression with nucleotide sequence, and the nucleotide sequence has
Under strict conditions with SEQ ID NO:The complementary series of nucleotide sequence hybridization shown in 2.
Embodiment three：
Present embodiments provide a kind of recombinant protein comprising destination protein and the polypeptide that can be inserted into the destination protein
Carrier, and the albumen that the destination protein is the separation in embodiment two；Preferably, the peptide carrier is prokaryotic expression carrier
Or carrier for expression of eukaryon, it is plasmid, bacteriophage etc. to include but not limited to.
Example IV：
Present embodiments provide a kind of polynucleotides, which is characterized in that it is used to encode such as the recombination egg in embodiment three
In vain.
Embodiment five：
Present embodiments provide a kind of carrier comprising more described in the nucleic acid molecules or example IV described in embodiment one
Nucleotide；Preferably, the carrier includes plasmid；And provide a kind of nucleic acid molecules including in embodiment one and/or above-mentioned
The host cell of carrier, and the host cell includes fish cell, particularly preferably teleost cell.
The building process of the carrier includes：
S1, design of primers：
It is sequenced by zebra fish transcript profile, obtains the nucleotide sequence of nox1；Specific primer is designed on this basis,
Including following sense primer and downstream primer；
Wherein, the base sequence of sense primer is：
5’-GTGAAGCTT(the local AAGCTT of underscore is the enzyme of Hind III to ACCATGGGCAACTGGATCATCAA-3 '
Enzyme site)
The base sequence of above-mentioned downstream primer is：
5’-GCAGGTACC(the local GGTACC of underscore is the digestion position of KPN I to AAGTTCTCTTTGTTGAAGTAA-3 '
Point)
S2, PCR amplification：
It is set to be denaturalized unwinding using 95 DEG C of high temperature using wild-type zebrafish cDNA as template, then temperature is down to 55 DEG C and is moved back
Fire makes above-mentioned primer be combined to form the conjugate of DNA profiling and primer with the single stranded DNA of template, is then warming up to 72 DEG C again, makes
Conjugate is under the action of Taq enzyme, and using target sequence as template, dNTP is raw material, according to base pairing and semi-conservative replicating principle
It is replicated, is repeated the above process, the open reading frame of a large amount of zebra fish nox1 is obtained with amplification；Then to target fragment and
Carrier carries out digestion with restriction enzyme Hind III and I enzymes of KPN simultaneously, and the nox1 segments cut are connected in p3xFLAG-CMV-
On 14 expression vectors；
S3, sequencing：
Recombinant plasmid is mixed with host cell first, it is made to be adhered to host cell surface, then uses 42 DEG C of temperature
Thermal shock 45s is spent, to promote host cell adequately to absorb recombinant plasmid, non-selection fluid nutrient medium is then added (without any
Antibiotic) about 1h cultures are carried out, then regathering host cell keeps it raw on Selective solid culture medium (containing antibiotic)
It is long, thus successfully screen the host cell of energy recombinant expression；
In the present embodiment, the host cell used is Escherichia coli TOP10.Since Escherichia coli breeding is fast, suitable
Under the conditions of the breeding cycle be only 20 minutes or so, and common plasmid can be proliferated in Escherichia coli to hundreds of copies.Cause
This can make the recombinant plasmid comprising target fragment a large amount of in a short time by cultivating the Escherichia coli after converting successfully
Amplification.
It selects three clones at random in bacterium colony and send sequencing, it is found that three clones are identical sequence and identification is correct, as
Zebra fish nox1_tv1, nucleic acid sequence such as SEQ ID NO：Shown in 1；The electrophoretogram of PCR amplification is as shown in Figure 1, therefrom can be with
Find out, amplifying nucleic acid sequence is identical as the nucleic acid size of zebra fish nox1_tv1 is estimated, and illustrates zebra fish nox1_tv1 recombination matter
Grain is built successfully.
Expression process of the carrier in host cell include：
S11, destination protein is collected：
By in the nox1_tv1 Transfected Recombinant Plasmids to fish cell of above-mentioned structure, total protein of cell is extracted after transfecting 48h；
S12, protein electrophoresis：
The total protein of cell of 20ul is added per hole for the PAGE gel for preparing 12%, first uses 80v electrophoresis 30min, then use
120v electrophoresis 90min；
S13, transferring film：
Destination protein is transferred on pvdf membrane, 80v voltage stabilizing transferring film 45min, transferring film success after with 5% skimmed milk power
(being prepared with TBST) close 1h, then plus the anti-monoclonal FLAG tag antibodies of mouse (Sigma companies, 1:5000 dilutions) 4 DEG C be incubated
Overnight, then film is washed with TBST 3 times, each 5min.Be eventually adding corresponding secondary antibody (ThermoFisher companies, 1:5000),
It is incubated after 1h and washes film 3 times with TBST, each 5min.
S14, colour developing：
ECL luminescent solutions are added, and gel imaging analysis, experimental result are as shown in Figure 2；By attached drawing 2 it is found that negative control is empty
Plasmid group does not have signal representation, target gene nox1_tv1 that can preferably be expressed in fish EPC cells.
Embodiment six：
Present embodiments provide a kind of pharmaceutical composition comprising the recombinant protein in embodiment three, and provide one
The purposes of kind aforementioned pharmaceutical compositions in medicine preparation, the drug is for inhibiting to include huichun viremia virus (SVCV)
Virus infection.
Specifically, the present invention examines table of the nox1_tv1 recombinant plasmids in the fish body infected by SVCV by qRT-PCR
Up to situation, concrete operations are as follows：
By microinjection by the embryo of the nox1_tv1 recombinant plasmids microinjection provided in embodiment five to zebra fish
Be used as test group in tire, at the same by the embryo of empty plasmid microinjection to zebra fish as a control group；Wait for that zebra fish juvenile fish is complete
It hatches entirely after coming, test group and control group is subjected to immersion infection, infection a concentration of 2 × 10 respectively6Pfu (plaque shapes
At unit)/ml, every group of 3 repetitions, every group of 30 tail fishes so that the juvenile fish of test group and control group infects SVCV；Feeling
The for 24 hours and 48h after dye, and every group takes about 20 tail fishes respectively, and after being washed with PBS, Trizol homogenate is added；Then 2 groups of samples are extracted
RNA, after reverse transcription is at cDNA with qRT-PCR (real time fluorescent quantitative) in real time detection and quantitative amount of product, experimental result is such as
Shown in table 1 and Fig. 3.
Table 1 infect SVCV after for 24 hours with nox1_tv1 expression quantity when 48h
By table 1 with Fig. 3 it is found that compared with control group, in test group, the (after fertilizations for 24 hours of nox1_tv1 after infection
5 days) and 48h (after fertilization the 6th day) raised 4 times and 5 times respectively.It follows that nox1_tv1 is infecting spring viremia
It can preferably be expressed in the fish body of virus.
Further, the present invention examines nox1_tv1 recombinant plasmids mistake in the fish body infected by SVCV by qRT-PCR
The influence to virus multiplication is expressed, concrete operations are as follows：
By microinjection by the embryo of the nox1_tv1 recombinant plasmids microinjection provided in embodiment five to zebra fish
Be used as test group in tire, at the same by the embryo of empty plasmid microinjection to zebra fish as a control group；Wait for that zebra fish juvenile fish is complete
It hatches entirely after coming, test group and control group is subjected to immersion infection, infection a concentration of 2 × 10 respectively6Pfu (plaque shapes
At unit)/ml, every group of 3 repetitions, every group of 30 tail fishes so that the juvenile fish of test group and control group infects SVCV；Feeling
The for 24 hours and 48h after dye, and every group takes about 20 tail fishes respectively, and after being washed with PBS, Trizol homogenate is added；Then 2 groups of samples are extracted
RNA, after reverse transcription is at cDNA with qRT-PCR (real time fluorescent quantitative) in real time detection SVCV N protein (SVCV-N) and G eggs
The expression of (SVCV-G) in vain, experimental result is as shown in table 2 and Fig. 4-5.
Table 2 infects the expression quantity with SVCV-N when 48h and SVCV-G for 24 hours after SVCV
By table 2 and Fig. 4,5 it is found that compared with the control group, the SVCV-N of test group is after infection for 24 hours (after fertilization the 5th day)
0.35 and 0.43 times has been lowered respectively with 48h (after fertilization the 6th day), and SVCV-G has lowered 0.38 respectively with 48h for 24 hours after infection
With 0.54 times.It can be seen that overexpression of the nox1_tv1 recombinant plasmids in fish body can significantly inhibit SVCV viruses in vivo
Proliferation.
In addition, the present embodiment is also overexpressed in fish body to virus by the method that the death rate counts to detect nox1_tv1
The influence of metainfective juvenile fish survival, concrete operations are as follows：
By microinjection by the embryo of the nox1_tv1 recombinant plasmids microinjection provided in embodiment five to zebra fish
Be used as test group in tire, at the same by the embryo of empty plasmid FLAG microinjections to zebra fish as a control group；Wait for zebra fish children
After fish hatches completely, test group and control group are subjected to immersion infection, infection a concentration of 2 × 10 respectively6Pfu is (empty
Spot forms unit)/ml, every group of 3 repetitions, every group of 35 tail fishes so that the juvenile fish of test group and control group infects SVCV.
The dead quantity is counted daily, draws survival curve after counting 7 days altogether, experimental result is as shown in table 3 and Fig. 6.
Table 3 infects the survival rate after SVCV
As shown in table 3 and Fig. 6, overexpressions of the nox1_tv1 in fish can significantly improve fish body and feel in virus experimental result
Survival rate during dye.(control group survival rate is 56.19%) is compared with control group, nox1_tv1 injections group is in SVCV virus senses
Survival rate after dye improves 27.62% (nox1_tv1 injection group survival rates are 83.81%).
Meanwhile the present embodiment is overexpressed in fish body to antiviral base by the method for qRT-PCR to detect nox1_tv1
The influence of cause, concrete operations are as follows：
By microinjection by the embryo of the nox1_tv1 recombinant plasmids microinjection provided in embodiment five to zebra fish
Be used as test group in tire, at the same by the embryo of empty plasmid microinjection to zebra fish as a control group；Wait for that zebra fish juvenile fish is complete
It hatches entirely after coming, test group and control group is subjected to immersion infection, infection a concentration of 2 × 10 respectively6Pfu (plaque shapes
At unit)/ml, every group of 3 repetitions, every group of 30 tail fishes so that the juvenile fish of test group and control group infects SVCV；Feeling
1d (1dph) and 2d (2dph) after dye, every group takes about 30 tail fishes respectively, and after being washed with PBS, Trizol homogenate is added；Extraction
The RNA of 2 groups of samples detects the expression of IFN1 and MXA genes in real time after reverse transcription is at cDNA with qRT-PCR, and experimental result is such as
Shown in table 4 and Fig. 7-8.
Table 4 infect SVCV after 1d and 2d when IFN1 and MXA expression quantity
By table 4 and attached drawing 7-8 it is found that being compared with injection empty plasmid group, overexpression energy of the nox1_tv1 in zebra fish body
The significantly transcription of induction Antiviral related gene.IFN1 1dph (after fertilization the 5th day) and 2dph (after fertilization the 6th day) respectively on
Adjust 1.9 and 4.8 times；MXA raises 2.3 and 3.1 times respectively in 1dph and 2dph.
In conclusion the nucleic acid molecules of the present invention can quickly and efficiently inhibit duplication of the virus in fish body, effectively
The immunity of host is improved, and its overexpression in fish body can significantly improve survival rate of the fish body during SVCV infects.
The foregoing is merely presently preferred embodiments of the present invention, is not intended to limit the invention, it is all the present invention spirit and
Within principle, any modification, equivalent replacement, improvement and so on should all be included in the protection scope of the present invention.
Claims (12)
1. a kind of nucleic acid molecules, which is characterized in that including：(a) there is SEQ ID NO:Nucleotide sequence shown in 1；Or
(b) there is SEQ ID NO:Nucleotide sequence shown in 1 can variant sequence thereof, and it is described can variant sequence thereof and SEQ ID NO:
Nucleotide sequence shown in 1 has at least 80% homology；Or
It (c) under strict conditions can be with the nucleotide sequence of the nucleotide sequence hybridization limited in (a)；Or
(d) there is SEQ ID NO:Nucleotide sequence shown in 2；Or
(e) there is SEQ ID NO:Nucleotide sequence shown in 2 can variant sequence thereof, and it is described can variant sequence thereof and SEQ ID NO:
Nucleotide sequence shown in 2 has at least 80% homology；Or
It (f) under strict conditions can be with the nucleotide sequence of the nucleotide sequence hybridization limited in (d)；Or
(g) coding has SEQ ID NO:The nucleotide sequence of the polypeptide of amino acid sequence shown in 3；Or
(h) nucleotide sequence of encoding amino acid sequence, and the amino acid sequence is SEQ ID NO:Amino acid sequence shown in 2
The polypeptide sequence formed after replacing, missing or adding one or more amino acid；Or
(i) coding has SEQ ID NO:The nucleotide sequence of the polypeptide of amino acid sequence shown in 4；Or
(j) nucleotide sequence of encoding amino acid sequence, and the amino acid sequence is SEQ ID NO:Amino acid sequence shown in 4
The polypeptide sequence formed after replacing, missing or adding one or more amino acid.
2. a kind of protein of separation, which is characterized in that including：(a) there is SEQ ID NO:The albumen of amino acid sequence shown in 3
Matter；Or
(b) there is SEQ ID NO:Amino acid sequence shown in 3 is formed after replacing, missing or adding one or more amino acid
Sequence protein；Or
(c) by with SEQ ID NO:The protein that the nucleic acid molecules expression of nucleotide sequence shown in 1 generates；Or
(d) protein generated by the nucleic acid molecules expression with nucleotide sequence, and the nucleotide sequence has tight
Under the conditions of lattice with SEQ ID NO:The complementary series of nucleotide sequence hybridization shown in 1.
(e) there is SEQ ID NO:The protein of amino acid sequence shown in 4；Or
(f) there is SEQ ID NO:Amino acid sequence shown in 4 is formed after replacing, missing or adding one or more amino acid
Sequence protein；Or
(g) by with SEQ ID NO:The protein that the nucleic acid molecules expression of nucleotide sequence shown in 2 generates；Or
(h) protein generated by the nucleic acid molecules expression with nucleotide sequence, and the nucleotide sequence has tight
Under the conditions of lattice with SEQ ID NO:The complementary series of nucleotide sequence hybridization shown in 2.
3. a kind of recombinant protein, which is characterized in that the peptide carrier including destination protein and the pluggable destination protein, and
The destination protein is the albumen of the separation in claim 2.
4. recombinant protein as claimed in claim 3, which is characterized in that the peptide carrier is prokaryotic expression carrier or eukaryon table
Up to carrier.
5. a kind of polynucleotides, which is characterized in that it is used to encode recombinant protein as described in claim 3 or 4.
6. a kind of carrier, which is characterized in that including nucleic acid molecules as described in claim 1 or as claimed in claim 5 more
Nucleotide.
7. carrier as claimed in claim 4, which is characterized in that the carrier includes plasmid.
8. a kind of host cell, which is characterized in that including nucleic acid molecules as described in claim 1 and/or such as claim 6 or
Carrier described in 7.
9. host cell as claimed in claim 8, which is characterized in that the host cell includes fish cell.
10. a kind of pharmaceutical composition, which is characterized in that including recombinant protein as claimed in claim 3.
11. the purposes of pharmaceutical composition according to any one of claims 10 in medicine preparation, the drug is for inhibiting viral infection.
12. purposes as claimed in claim 11, which is characterized in that the virus includes huichun viremia virus.
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Citations (3)
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|---|---|---|---|---|
| US20140221457A1 (en) * | 2013-01-24 | 2014-08-07 | University Of Central Florida Research Foundation, Inc. | Ameliorating oxidative stress in neurodegenerative disease via nox1 targeting |
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| US20140221457A1 (en) * | 2013-01-24 | 2014-08-07 | University Of Central Florida Research Foundation, Inc. | Ameliorating oxidative stress in neurodegenerative disease via nox1 targeting |
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| CN104630348A (en) * | 2015-01-20 | 2015-05-20 | 湖州市第一人民医院 | RT-PCR primers for detecting transcriptional level of NOX1 mRNA in serum as well as kit and method for evaluating concurrent intestinal cancer susceptibility of hyperglycemia population |
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