CN102524177B - Method for constructing HRN/gpt delta transgenic mouse model - Google Patents
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Abstract
The invention discloses a method for constructing a liver cytochrome P450 enzyme function deficiency type gpt delta transgenic mouse model. The mouse model provided by the invention has all characteristics of a liver cytochrome P450 reductase-removed mouse and a gpt delta transgenic mouse, the research on the relationship between compound metabolism and mutagenesis can also be carried out, and the mouse model is especially applicable to researching compounds making extrahepatic tissue organs activated by liver CYP metabolism mutate.
Description
Technical field
The present invention relates to the construction method of animal model, in particular to a kind of liver cytochrome P 450 enzyme functional defect type gpt delta(HRN/gpt delta) construction method of transgene mouse model.
Background technology
The organism metabolism process both can have been detoxified to xenobiontics also can increase poison, and closely related to the carcinogenesis of body with compound.Liver cytochrome P 450 enzyme (being abbreviated as CYP) plays an important role in the metabolic process of body to exogenous compounds, but it is very limited to can be used for disclosing the research tool of its carcinogenesis.
At present, research liver P450 enzyme mainly concentrates on the experiment in vitro the metabolism of exogenous compounds, comprises the application of liver microsomes, primary hepatocyte, each hypotype high expressing cell strain of CYPs and recombinase.Although these in-vitro methods have very big using value, but, because the absorption approach difference of exogenous compounds, the existence of organic the moon (sun) ion transport body in the tissue, the existence of complicated factors such as the outer CYPs effect of kidney scavenging action and liver, make above-mentioned experiment in vitro still be difficult to accurately predicting exogenous compounds actual metabolic pathway (Friedberg T in vivo, Pritchard MP, Bandera M, Hanlon SP, Yao D, McLaughlin LA, Ding S, Burchell B, Wolf CR.Merits and limitations of recombinant models for the study of human P450-mediated drug metabolism and toxicity:an intralaboratory comparison.Drug Metab Rev.1999.31 (2): 523-44.).In the metabolism research, have a plurality of P450 enzyme hypotype knock-out mice models at present in vivo, these models have been widely used for studying specificity P450 enzyme hypotype in the metabolism of exogenous compounds and the effect in the toxicity.Yet CYP has multiple hypotype usually, and each hypotype function class seemingly, and substrate exists and intersects, and makes the gene knockout animal model of single hypotype be difficult to accurately estimate the influence of P450 enzyme to exogenous compounds toxicity.Therefore, the model that makes all CYPs all lose function will be a very useful instrument.NADPH-cytochrome P450 reductase (CPR/POR) is the oxidoreduction partner of all microsome P450 enzymes, it provides first required electronics of P450 oxydasis reduction reaction, based on this principle, people such as Jun Gu developed a liver specificity P450 reductase-Cpr gene knock-out mice model (
HEpatic
REductase
NUll, HRN is designated hereinafter simply as liver specificity CPR KO mouse model), the disappearance of this gene expression will suppress activity (the Wu L of all liver P450 enzymes, Gu J, Weng Y, Kluetzman K, Swiatek P, Behr M, Zhang QY, Zhuo X, Xie Q, Ding X.Conditional knockout of the mouse NADPH-cytochrome p450reductase gene.Genesis.2003Aug; 36 (4): 177-81.).This model utilizes the Cre/LoxP system to knock out (conditional knock) by condition and obtains, mouse liver Cpr gene in maturation is knocked out (being born after 2 months) gradually, finally make liver P450 enzymic activity lose, and its hetero-organization is unaffected.
Most carcinogenic substances have genetoxic, promptly damage genetic material (DNA or chromosome), cause gene mutation and then carcinogenic.Genotoxic evaluation is the effective ways of prediction carcinogenicity, and the object of its detection comprises the change of gene mutation, chromosome damage, reorganization and number that dna damage and dna damage cause etc.Estimate employing standard combination test usually and carry out, as Salmonella reversion test, mammalian cell gene mutation test and the transgenic animal sudden change detection test etc. of the Using Comet Assay of estimating dna damage, the micronucleus test of estimating chromosome damage and chromosomal aberration test, evaluation gene mutation.Compare with experiment in vitro, vivo experiment method false positive incidence is lower, and has the advantage of considering absorption, metabolism, distribution and the drainage relevant with the human body application, so the application of experiment in the body comes into one's own day by day.Existing In vivo assay Cells is less, the measurable carcinogenicity of micronucleus test in the classical body, but this method is only estimated marrow and can not be predicted the toxicity target organ.Transgenic animal are very important research tools, can detect any required intraorganic gene mutation, the mechanism analysis that can also suddenly change.
Transgenic animal sudden change detection model always mainly contains (J A Gossen such as Muta Mouse and Big Blue Mouse/Rat, W J de Leeuw, C H Tan, E C Zwarthoff, F Berends, P H Lohman, D L Knook, and J Vijg.Efficient rescue of integrated shuttle vectors from transgenic mice:a model for studying mutations in vivo, Proc.Natl.Acad.Sci.U.S.A.1989,86:7971-7975.Kohler SW, Provost GS, Kretz PL, Fieck A, Sorge JA, Short JM.The use of transgenic mice for short-term, in vivo mutagenicity testing.Genet Anal Tech Appl.1990Dec; 7 (8): 212-8.).But the common defects that above-mentioned model has be can only test point the deletion mutation of sudden change and DNA small fragment, and the deletion mutation of fragment just is difficult to be detected greatly.1996, people such as Nohmi set up new transgenic animal model gpt delta transgenic mice (Nohmi T, Katoh M, Suzuki H, Matsui M, Yamada M, Watanabe M, Suzuki M, Horiya N, Ueda O, Shibuya T, Ikeda H, Sofuni T.A new transgenic mouse mutagenesis test system using Spi-and6-thioguanine selections.Environ Mol Mutagen.1996; 28 (4): 465-70.), the characteristics of this model are the sudden changes that not only can detect bases such as comprising point mutation, frameshift mutation, base replacement in the Different Organs of mouse, can also detect dna fragmentation and lack on a large scale.In addition, the length of gpt gene is 456bp, helps further by order-checking mechanism analysis being carried out in sudden change.
Determine that the relation between mutagenesis detects in liver cytochrome P 450 and the body has great significance to the genetic predisposition prediction and the prevention of compound.In two kinds of mouse models of this that has, liver specificity CPR KO mouse model can not carry out vivo mutations and detect at present, detects though gpt delta transgenic mice can be done vivo mutations, can't investigate liver cytochrome P 450 role therein.Therefore, the animal model that has only foundation to possess two specific characters simultaneously just can address this problem.
Summary of the invention
At deficiency of the prior art, the inventor is devoted to study the animal model that possesses two specific characters simultaneously.Thus, the present invention is hybridized two kinds of animal pattern gpt delta transgenic mices using with the liver specificity CPR KO mouse with identical C57BL/6J genetic background, by detecting genotype, select required filial generation animal to carry out mating repeatedly, finally obtained the gpt delta transgenic mice new model of liver specificity CPR deficiency, be liver P450 enzyme functional defect type gpt delta transgenic mice (
HEpatic
REductase
NUll gpt delta transgenic mice abbreviates HRN/gpt delta transgenic mice as).And it is unsuccessful that the present invention has also overcome hybridization in repetition test, filial generation can not survival or genotype be not inconsistent with expection, and make the hybridization technical barrier that afterwards each gene of mouse can genetic stability.
Therefore, an object of the present invention is to provide a kind of liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model.
Another object of the present invention provides a kind of construction method of liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model.
A further object of the present invention provides the application in the relation between research liver cell P450 enzyme and compound mutagenesis of this model.
For achieving the above object, the invention provides a kind of liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model.
According to another object of the present invention, the invention provides a kind of method that makes up above-mentioned model, this method may further comprise the steps:
(1) selects for use liver specificity CPR KO mouse and gpt delta transgenic mice to hybridize respectively, obtain F1 generation: cpr
Loxp+/-Cre
+/-Gpt
+/-And cpr
Loxp+/-Cre
-/-Gpt
+/-Mouse genotypes.Because two kinds of mouse genetic background unanimities all are C57BL/6J strain mouse, so there is not the genetic background difference problem in generation mice.
(2) with the cpr of F1 in generation
Loxp+/-Cre
+/-Gpt
+/-Genotypic female mouse and male mouse hybridize again, obtain F2 generation: cpr
Loxp+ /+Cre
+/-Gpt
+ /+And cpr
Loxp+ /+Cre
-/-Gpt
+ /+Mouse genotypes.
(3) with the cpr of F2 in generation
Loxp+ /+Cre
+/-Gpt
+ /+Male mouse of genotype and cpr
Loxp+ /+Cre
-/-Gpt
+ /+The female mouse of genotype carries out mating, produces the more F3 cpr in generation
Loxp+ /+Cre
+/-Gpt
+ /+And cpr
Loxp+ /+Cre
-/-Gpt
+ /+Mouse genotypes.
(4) with the cpr of F3 in generation
Loxp+ /+Cre
+/-Gpt
+ /+Male mouse of genotype and cpr
Loxp+ /+Cre
-/-Gpt
+ /+The female mouse of genotype is the liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model that can normally be bred of post-coitum again.
Above-mentioned steps all needs to utilize the specific gene amplimer to carry out genotype identification to the mouse that is obtained.
According to another object of the present invention, the invention provides the application in the relation between research liver cell P450 enzyme and compound mutagenesis of above-mentioned mouse model.
The beneficial effect that liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model of the present invention is compared with conventional art is as follows:
At first, compared with prior art, liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model of the present invention is by obtaining liver specificity CPR KO mouse and gpt delta transgenic mice behind a series of hybridization and genotype screening, the feature that it has two kinds of parental generation models concurrently is the indispensable technological means of relation between research liver P450 enzyme and the compound mutagenesis.
Secondly, new model of the present invention also possesses whole application of two kinds of parental generation models:
A. the application that has liver specificity CPR KO mouse aspect: whether compound passes through metabolism and the effect of liver CYP; The effect of CYP in the compound metabolism in outer its hetero-organization internal organs of liver; CYP totally different in endogenous and exogenous compounds metabolism in liver microsomes and the mitochondria; Liver HO enzyme function changes the relevant metabolic alterations in back; Also can be used for research can CYP metabolism in liver, but in the extrahepatic tissue internal organs toxigenous compound; Be used to study the compound that hepatotoxicity wind agitation strengthens after the liver CYP metabolism.
B. the application that has gpt delta transgenic mice aspect: because λ EG10DNA is present in the genome of mouse, therefore can do detection in Gene Mutation and analysis to the internal organs of organizing of mouse whole body, but test point sudden change and deletion mutation; And compound is to the comparative studies of the mutagenesis of different tissues organ.
Once more, this new model not only has liver cytochrome P 450 reductase knock-out mice and all features of gpt delta transgenic mice, but also can carry out the research that concerns between compound metabolism and the mutagenesis, be particularly suitable for the research that those can be formed the compound of sudden change by liver CYP metabolism activation extrahepatic tissue organ.
In addition, in the process that new model makes up, it is unsuccessful that the present invention has also overcome hybridization, filial generation can not survival or the difficult problem that is not inconsistent of genotype and expection, this is because liver specificity CPR KO mouse and gpt delta transgenic mice all are C57BL/6J strain mouse, genetic background is more consistent, so the hybridization chance of success is bigger; Liver specificity CPR KO mouse and gpt delta transgenic mice fertility are normal, non-interaction action between cpr gene knockout mechanism and the λ EG10 sequence in the new mouse genotypes that the hybridization back forms, so generation mice not only can be survived but also fertility is normal.And hybridization many different mouse genotypes may occur afterwards because liver specificity CPR KO mouse is with gpt delta transgenic mice, and genotype screening process of the present invention can effectively solve this difficult problem.Further, the inventor is at cpr
LoxpGene, cre gene and gpt gene can the basis of genetic stability on, hybridize mice obtains cpr
LoxpNormally breed again after gene and the sub-mouse of gpt gene pure, so just guarantee the cpr in the new model mouse
LoxpGene, cre gene and gpt gene be genetic stability in each generation.
Description of drawings
Fig. 1 is the flow chart that liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model makes up.
Fig. 2 be F1 for the genotypic PCR qualification result of mouse, A is the mouse that the cre gene is arranged, wherein, cpr
LoxpGenetic heterozygosis expands and 500bp and 600bp two bands, and the cre gene expands and 300bp one band, gpt genetic heterozygosis expand 360bp(F-R1) and 967bp(F-R2) two bands; Be no cre dna murine among the B, wherein, cpr
LoxpGenetic heterozygosis expands and 500bp and 600bp two bands, gpt genetic heterozygosis expand 360bp(F-R1) and 967bp(F-R2) two bands.
Fig. 3 be F2 for the genotypic PCR qualification result of mouse, A is the mouse that the cre gene is arranged, wherein, cpr
LoxpGene pure expands and 600bp one band, and the cre gene expands and 300bp one band, gpt gene pure expand 967bp(F-R2) band; B is no cre dna murine, cpr
LoxpGene pure expands and 600bp one band, gpt gene pure expand 967bp(F-R2) band.
Fig. 4 is according to main organs qualification result in the gpt delta transgenic mice of the liver specificity CPR deficiency in F2 generation in one embodiment of the present invention.A is cpr
LoxpGene and cre gene, B are the gpt gene, and wherein 967bp is the PCR result of primers F-R2 of using in the genotype identification, and 600bp is primer gpt-F and gpt-R qualification result; L: liver; K, kidney; St, stomach; B, bladder; Sp, spleen.
Embodiment
The present invention is further elaborated below in conjunction with embodiment, and following embodiment is only described the present invention by way of example.Clearly, those of ordinary skills can carry out various accommodations and modification to the present invention in scope of the present invention and essence.Need be appreciated that, this invention is intended to be encompassed in the flexible and modification that comprises in the appended claims.
The inventor is extensive studies through going deep into, make up a kind of inheritance stability, liver cytochrome P 450 enzyme functional defect type gpt delta transgene mouse model that phenotype is stable, be specially: selected for use the male liver specificity CPR of parental generation KO mouse and the female gpt delta of parental generation transgenic mice to hybridize (F0 generation) respectively.With the cpr of F1 in generation
Loxp+/-Cre
+/-Gpt
+/-Genotypic female mouse and male mouse hybridize and obtain F2 for cpr
Loxp+ /+Cre
+/-Gpt
+ /+And cpr
Loxp+ /+Cre
-/-Gpt
+ /+Mouse genotypes is genotypic mouse required for the present invention, a large amount of screenings of this process need.Then, with the cpr of F2 in generation
Loxp+ /+Cre
+/-Gpt
+ /+Male mouse and cpr
Loxp+ /+Cre
-/-Gpt
+ /+Female mouse carries out expanding propagation by mating, thereby produces the more F3 cpr in generation
Loxp+ /+Cre
+/-Gpt
+ /+And cpr
Loxp+ /+Cre
-/-Gpt
+ /+Mouse genotypes.With the F3 cpr in generation
Loxp+ /+Cre
+/-Gpt
+ /+Male mouse and cpr
Loxp+ /+Cre
-/-Gpt
+ /+Female mouse is post-coitum again, and F4 is carried out the identified gene type for sub-mouse, but the result shows three kinds of equal genetic stabilities of gene in the sub-mouse of gained, promptly illustrates from F3 for can normally breeding.Entire making process all needs to utilize the specific gene amplimer to carry out genotype identification.
Specific implementation process:
1.F0 generation hybridization:
The male liver specificity CPR of parental generation KO mouse and the female gpt delta of parental generation transgenic mice are hybridized, and produce F1 generation.Cut toe identified gene type at F1 4 weeks for generation mice birth back.
1.1 extraction genomic DNA:
The toe of cutting is put into the Ep pipe of 1.5mL, add the DNA digestion buffer solution that 0.4mL contains the 0.4mg/mL Proteinase K, put into water-bath then, 55 ℃ of digested overnight (digestion time is more than 16h).Next day, treat mouse toe digestion fully after, in each Ep pipe, add 0.4mL phenol chloroform (pH8.0), turn upside down and mix 50-60 time, 15,000g4 ℃ centrifugal 20 minutes.Get 300 μ L supernatants and insert in other 1 new 1.5mL Ep pipe,, put upside down gently up and down and mix 5-10 time to wherein adding 750 μ L ice absolute ethyl alcohol, 12, centrifugal 5 minutes of 000rpm.Abandon supernatant, add ice 75% ethanol 200 μ L and wash 2 times.The Ep pipe that will contain genomic DNA tips upside down on blotting paper last 30 minute, dry DNA.Add 80～100 μ L TE buffer solutions in the Ep pipe, room temperature was placed 2 hours, and DNA is fully dissolved.
1.2PCR detection genotype:
Be PCR with the genome that extracts, detect F1 for the mouse genotype.
1.2.1 primer sequence and product size:
Cpr
LoxpGene primer:
Upstream primer 1(F): 5 '-TACAATGGACCAGGCTCTGC-3 '
Downstream primer 2(R): 5 '-AAGAGGGACAAAGAGCACC-3 '
The PCR product: isozygoty, 600bp(contains loxp), a band; Heterozygosis 500bp(does not contain loxp) and 600bp(contain loxp) each band.
The Cre gene primer:
Upstream primer 3(F): 5 '-GGATTT CCGTCTCTGGTGTAGC-3 ' downstream primer 4(R): 5 '-CATTGCCCCTGTTTCACTATCC-3 '
PCR product: 300bp, a band.
The gpt gene primer:
Upstream primer 5(F): 5 '-GTTGTACTTCCAACCATGCCAAAG-3 '
Downstream primer 6(R1): 5 '-CAGAAATCATTCCAGGTCCTTGC-3 '
Downstream primer 7(R2): 5 '-GTTCATCTGCTTTATGGGCAAGAG-3 '
Upstream primer 8(gpt-F): 5 '-GCGCAACCTATTTTCCCCTCGA-3 '
Downstream primer 9(gpt-R): 5 '-TGGAAACTATTGTAACCCGCCTG-3 '
PCR product: homozygote, the PCR product of 967bp(primers F and primer R1), a band; Heterozygote is the PCR product of 360bp(primers F and primer R1) and the PCR product of 967bp(primers F and primer R2), two bands; Wild type has only the PCR product of 360bp(primers F and primer R1) band.Gpt-F and gpt-R product are 600bp, mainly identify the gpt gene.
1.2.2PCR program setting:
Cpr
LoxpWith Cre gene PCR program setting: 94 ℃ of pre-sex change 3 minutes, 94 ℃ 30 seconds, 64 ℃ 30 seconds, 72 ℃ 30 seconds, totally 36 circulations, 72 ℃ were extended 4 ℃ of preservations 7 minutes then.
Gpt gene PCR program setting: 94 ℃ of pre-sex change 4.5 minutes, 94 ℃ 30 seconds, 58 ℃ 30 seconds, 72 ℃ 1 minute, totally 30 circulations, 72 ℃ were extended 4 ℃ of preservations 5 minutes then.
1.2.3PCR product is identified:
Getting 10 μ L PCR products after PCR finishes identifies in 1.5% agarose gel electrophoresis.Amplification is seen shown in Figure 1, and A shows the genotype of cre dna murine, and wherein, the 1st swimming lane is 100bpDNA ladder Marker, and the 2nd swimming lane is cpr
LoxpGenetic heterozygosis, 500bp and 600bp, the 3rd swimming lane are the cre gene, 300bp, the 4th, 5 swimming lanes are the sub-360bp(F-R1 of gpt genetic heterozygosis) and 967bp(F-R2).B shows the genotype of no cre gene (300bp) mouse among Fig. 1, and wherein, the 1st swimming lane is 100bp DNA ladder Marker, and the 2nd swimming lane is cpr
LoxpGenetic heterozygosis, 500bp and 600bp, the 4th, 5 swimming lanes are the sub-360bp(F-R1 of gpt genetic heterozygosis) and 967bp(F-R2).
The F1 that the hybridization back forms has two kinds for genotype: cpr
Loxp+/-Cre
+/-Gpt
+/-And cpr
Loxp+/-Cre
-/-Gpt
+/-
2.F1 generation hybridization:
With male and female cpr of F1 generation
Loxp+/-Cre
+/-Gpt
+/-The mouse mating, and to sub-mouse of F2 generation identified gene type in the manner described above.It is more that measurement result shows that F2 genotype occurs for sub-mouse, and screening also keeps cpr
Loxp+ /+Cre
+/-Gpt
+ /+And cpr
Loxp+ /+Cre
-/-Gpt
+ /+Mouse genotypes.
As shown in Figure 2, A shows the genotype of cre dna murine, and wherein, the 1st swimming lane is 100bp DNA ladder Marker, and the 2nd swimming lane is cpr
LoxpGene pure, 600bp, the 3rd swimming lane are the cre gene, 300bp, the 4th, 5 swimming lanes are gpt gene pure, no 360bp(F-R1) gene outcome, have only 967bp(F-R2).B shows the genotype of no cre gene (300bp) mouse among Fig. 2, and wherein, the 1st swimming lane is 100bp DNA ladder Marker, and the 2nd swimming lane is cpr
LoxpGene pure, 600bp, the 4th, 5 swimming lanes are gpt gene pure, no 360bp(F-R1) gene outcome, have only 967bp(F-R2).
3.F2 for expanding propagation:
With male cpr of F2 generation
Loxp+ /+Cre
+/-Gpt
+ /+Mouse and female cpr
Loxp+ /+Cre
-/-Gpt
+ /+The mouse mating, and to sub-mouse of F3 generation identified gene type in the manner described above.Because F2 is for cpr in the sub-mouse
Loxp+ /+Cre
+/-Gpt
+ /+Mouse and cpr
Loxp+ /+Cre
-/-Gpt
+ /+The mouse proportion is less, therefore need be with two kinds of mouse post-coitum expanding propagation.
4.F3 normal breeding of generation:
With male cpr of F3 generation
Loxp+ /+Cre
+/-Gpt
+ /+Mouse and female cpr
Loxp+ /+Cre
-/-Gpt
+ /+The mouse mating, and to sub-mouse of F4 generation identified gene type in the manner described above.The result shows, can normally breed from F3 generation, but three kinds of equal genetic stabilities of gene.
Sequence table
＜110〉Shanghai Pharmaceutical Inst., Chinese Academy of Sciences
＜120〉construction method of HRN/gpt delta transgene mouse model
<130> DI10-2039-XC68
<160> 9
<170> PatentIn version 3.3
<210> 1
<211> 20
<212> DNA
＜213〉artificial sequence
<400> 1
tacaatggac caggctctgc 20
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＜213〉artificial sequence
<400> 2
aagagggaca aagagcacc 19
<210> 3
<211> 22
<212> DNA
＜213〉artificial sequence
<400> 3
ggatttccgt ctctggtgta gc 22
<210> 4
<211> 22
<212> DNA
＜213〉artificial sequence
<400> 4
cattgcccct gtttcactat cc 22
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＜213〉artificial sequence
<400> 5
gttgtacttc caaccatgcc aaag 24
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<400> 6
cagaaatcat tccaggtcct tgc 23
<210> 7
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＜213〉artificial sequence
<400> 7
gttcatctgc tttatgggca agag 24
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<211> 22
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＜213〉artificial sequence
<400> 8
gcgcaaccta ttttcccctc ga 22
<210> 9
<211> 23
<212> DNA
＜213〉artificial sequence
<400> 9
tggaaactat tgtaacccgc ctg 23
Claims (4)
1. liver cytochrome P 450 enzyme functional defect type
GptThe construction method of delta transgene mouse model is characterized in that, may further comprise the steps:
Select for use respectively liver specificity CPR KO mouse and
GptThe delta transgenic mice is hybridized, and obtains F1 generation:
Cpr Loxp+/-
Cre +/- Gpt +/-With
Cpr Loxp+/-
Cre -/- Gpt +/-Mouse genotypes;
With F1 in generation
Cpr Loxp+/-
Cre +/- Gpt +/-Genotypic female mouse and male mouse hybridize again, obtain F2 generation:
Cpr Loxp+ /+
Cre +/- Gpt + /+With
Cpr Loxp+ /+
Cre -/- Gpt + /+Mouse genotypes;
With F2 in generation
Cpr Loxp+ /+
Cre +/- Gpt + /+The male mouse of genotype with
Cpr Loxp+ /+
Cre -/- Gpt + /+The female mouse of genotype carries out mating, produces more F3 generation
Cpr Loxp+ /+
Cre +/- Gpt + /+With
Cpr Loxp+ /+
Cre -/- Gpt + /+Mouse genotypes; And
With F3 in generation
Cpr Loxp+ /+
Cre +/- Gpt + /+The male mouse of genotype with
Cpr Loxp+ /+
Cre -/- Gpt + /+The female mouse of genotype post-coitum again can obtain the liver cytochrome P 450 enzyme functional defect type that can normally breed
GptThe delta transgene mouse model.
2. the method for claim 1 is characterized in that, described CPR KO mouse and
GptThe delta transgenic mice has identical C57BL/6J genetic background.
3. the method for claim 1 is characterized in that, described each step further comprises utilizes the specific gene amplimer to carry out genotype identification to the mouse that is obtained.
4. liver cytochrome P 450 enzyme functional defect type as claimed in claim 1
GptThe delta transgene mouse model is the application in the relation between research liver cell P450 enzyme and compound mutagenesis.
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