CN101342372B - Use of pancreatic lipase associated protein 1 - Google Patents
Use of pancreatic lipase associated protein 1 Download PDFInfo
- Publication number
- CN101342372B CN101342372B CN2007100435817A CN200710043581A CN101342372B CN 101342372 B CN101342372 B CN 101342372B CN 2007100435817 A CN2007100435817 A CN 2007100435817A CN 200710043581 A CN200710043581 A CN 200710043581A CN 101342372 B CN101342372 B CN 101342372B
- Authority
- CN
- China
- Prior art keywords
- plrp1
- gene
- mice
- obesity
- diabetes
- 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.)
- Active
Links
Images
Landscapes
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The present invention discloses an obesity/diabetes animal model, a method of setting up the obesity/diabetes animal model and applications of the obesity/diabetes animal model. In the obesity/diabetes mouse model, the PLRP1 gene is removed from the genome, or an exogenous gene is inserted into the PLRP1 gene, which leads to the inactivation of the PLRP1 gene. The genetic stability and phenotypic stability of the obesity/diabetes animal model can be used in the research of the physiological functions and pathologic functions of pancreatic lipase related proteins 1 in the body as well as in the screening and development of drugs for curing obesity and diabetes.
Description
Technical field
The present invention relates to medical science, materia medica and field of transgenic technology.Particularly, the present invention relates to a kind of obesity/diabetic mice model, set up the method for this obesity/diabetic mice model, and purposes that should obesity/diabetic mice model.More specifically, the present invention relates to utilize model organism research pancreatic lipase associated protein 1 physiology and pathology function in vivo, and find that further it can be used for fat and diagnosis of diabetes and treatment, and can be used for the screening and the development of fat and Remedies for diabetes.
Background technology
Lipase is a kind of water miscible enzyme, and the ester bond in some non-water soluble substance such as energy hydrolyzing triglyceride, phospholipid and cholesteryl ester plays important effect in the processes such as food digestion, fat absorption, equilibrium energy and plasma lipoprotein metabolism.According to function roughly the same and the homology on aminoacid sequence, the lipase family member comprises pancreatic lipase (pancreatic lipase at present, PL), lipoprotein lipase (lipoprotein lipase, LPL) and hepatic lipase (hepatic lipase, HL), pancreatic lipase associated protein 1 (Pancreatic lipase related proteins1, PLRP1), pancreatic lipase associated protein 2 (Pancreatic lipase related proteins2, PLRP2), phosphatidyl serine E.C. 3.1.1.32 (Phosphatidylserine-specific Phospholipase A1, PSPL A1) and the endothelium endogenous lipase (endothelial lipase, EL).
People's pancreatic lipase associated protein 1 (PLRP1) was delivered (Giller T in 1992 by the clone, BuchwaldP, Blum-Kaelin D, Hunziker W.Two novel human pancreatic lipase relatedproteins.hPLRP1and hPLRP2.Differences in colipase dependence and inlipase activity.J Biol Chem.1992Aug15; 267 (23): 16509-16.), the homology of its aminoacid sequence and people's pancreatic lipase sequence is 68%, has lineal homologous genes in mice.In pancreas liquid, can detect this albumen, show that it can be secreted into extracellular (De Caro J, Carriere F, Barboni P, Giller T, Verger R, De Caro A.Pancreatic lipase-relatedprotein1 (PLRP1) is present in the pancreatic juice of several species.Biochim Biophys Acta.1998Sep8; 1387 (1-2): 331-41.).
Although PLRP1 and PL have very high homology, but existingly studies show that natural PLRP1 does not have tangible esterase active (Payne RM, Sims HF, Jennens ML, Lowe ME.Rat pancreaticlipase and two related proteins:enzymatic properties and mRNAexpression during development.Am J Physiol.1994May; 266 (5Pt1): G914-21.).
In sum, at present not clear to the function of PLRP1 gene and the effect in integral body, there is not bibliographical information at the mice gene knockout research of PLRP1 yet.
The research of carrying out function unknown gene in the body has many methods, wherein the mice gene knochout technique provides very strong means for the function of research human gene in integral body, change by the phenotype of analyzing the genetic modification mice that obtains, just can obtain the function information of this gene; Another advantage of this research method be gene function and disease can be carried out related, thereby when obtaining gene function, also can obtain disease information and the disease animal model that this gene can be treated as potential drug or drug target.
On the other hand, because Developmental and Metabolic Disorder institute directly or the disease that causes indirectly, as obesity, diabetes and cardiovascular disease etc., has become the principal disease of current threat human health.With regard to diabetes, according to the WHO statistics, there were 3,000 ten thousand diabetes patients in the whole world in 1985, increased to 1.35 hundred million by 1997, and present global diabetic has 1.5 hundred million approximately, will reach 300,000,000 to global diabetics in 2025.And in the industrial city, the population of obesity has surpassed 10% of total population.The generation of Developmental and Metabolic Disorder is relevant with the deterioration of human life style's change and living environment, and also forming with the human gene simultaneously has substantial connection.
In order to research and develop metabolic diseases such as preventing and/or treating obesity, diabetes, hyperglycemia, this area presses for exploitation related gene kind and inheritance stability, obesity/diabetes animal model that phenotype is stable.
Summary of the invention
Purpose of the present invention just provides a kind of method that is used to set up inheritance stability, obesity/diabetic mice model that phenotype is stable, and the transgene mouse model that is used for this method.
Another object of the present invention provides the purposes of this obesity/diabetic mice model.
In a first aspect of the present invention, a kind of method that produces the non-human mammal of obesity/diabetes is provided, comprise step:
(a) by rejecting the PLRP1 gene or insert exogenous gene from genome in the PLRP1 gene, produce the embryonic stem cell of the non-human mammal of PLRP1 gene inactivation, wherein PLRP1 is a pancreatic lipase associated protein 1;
(b) with the embryonic stem cell regeneration non-human mammal in the step (a), the PLRP1 gene is owing to rejecting or inserting exogenous gene and inactivation in the genome of described non-human mammal.
In another preference, described step (b) is that the embryonic stem cell with step (a) is expelled in the segmentation cavity and is transplanted to the fallopian tube or the uterus of the non-human mammal of pseudo-fetus, produces the non-human mammal of PLRP1 gene inactivation then.
In another preference, described step (a) is to cause the PLRP1 inactivation by the exon of replacing the PLRP1 gene with the Neo gene.
In another preference, described method also comprises step:
(c) non-human mammal that step (b) is obtained is identified.
In another preference, comprise also that in step (b) non-human mammal that obtains is carried out copulation to be built and be, thereby obtain filial generation.
In another preference, described non-human mammal is mice, rat, rabbit, monkey.
In another preference, the non-human mammal in the step (b) isozygotys, and promptly two PLRP1 genes in the genome are all owing to rejecting or inserting exogenous gene and inactivation.
In a second aspect of the present invention, a kind of purposes of using the non-human mammal of said method acquisition of the present invention is provided, these non-human mammals are used as obesity/diabetes model; Or be used to screen and prevent and/or treat fat medicine; And/or be used to screen the medicine of prevention and/or diabetes.
In another preference, described non-human mammal is mice or rat.
In another preference, described non-human mammal isozygotys, and promptly two PLRP1 genes in the genome are all owing to rejecting or inserting exogenous gene and inactivation.
In a third aspect of the present invention, the purposes of a kind of pancreatic lipase associated protein 1 (PLRP1) or its encoding gene or its regulating and controlling sequence is provided, they are used to prepare the reagent of diagnosis obesity, diabetes and/or hyperglycemia by (a); (b) be used to prepare or screen that treatment is fat, the medicine of diabetes and/or hyperglycemia.
Should understand, technical characterictic above-mentioned necessity of the present invention and preferred and other technologies feature (comprising disclosed technical characterictic among the embodiment), constitute various technical scheme thereby can make up mutually, these technical schemes all constitute the application's disclosure.
Description of drawings
Fig. 1 has shown the structure of targeting vector and the strategy of practicing shooting.
Fig. 2 has shown the electrophoretogram of identifying three kinds of genotypic mices with PCR.
Fig. 3 has shown Western trace result, proves that this gene knockout mice do not express PLRP1 albumen.
Fig. 4 has shown that the fat content of gene knockout mice (KO) is greater than wild type (WT).Wherein * represents P＜0.05.
Fig. 5 has shown that the blood glucose value of gene knockout mice (KO) is greater than wild type (WT).Wherein * represents P＜0.05.
Fig. 6 has shown the aminoacid sequence of people PLRP1.
Fig. 7 has shown the aminoacid sequence of mice PLRP1.
Fig. 8 has shown the sequence alignment result of people PLRP1 and mice PLRP1.
The specific embodiment
The inventor is through deeply and extensive studies, set up a kind of inheritance stability, obesity/diabetes model that phenotype is stable, and it is because the PLRP1 gene is disallowable or insert exogenous gene and mice or other non-human mammals of inactivation in the PLRP1 gene.Animal pattern of the present invention is obesity and/or diabetes model.Can be used for the research of lipid metabolism and carbohydrate metabolism, and can be used to screen the medicine that prevents and/or treats obesity and/or diabetes, especially prevent and/or treat the medicine of obesity and/or diabetes by regulation and control PLRP1 gene.Finished the present invention on this basis.
Particularly, the inventor utilizes the dna homology recombinant technique, the gene of PLRP1 of will encoding in mouse embryo stem cell (ES cell) realize to be rejected, and by mice blastaea injection technique, the mouse ES cells of PLRP1 gene mutation is expelled in 3.5 days the mice segmentation cavity, obtained gomphosis mouse, gomphosis mouse and the copulation of normal wild type mice are bred, screening has obtained the heterozygote mice (PLRP1-/+) of PLRP1 gene mutation in the offspring, the homozygote mice by the mutual copulation of heterozygote mice being obtained the PLRP1 gene mutation (PLRP1-/-).
Result of study shows, PLRP1-/-mice can normally be born, grow and ripe, and can procreate.The diet of homozygote mice contrasts with wild type with motion and compares Non Apparent Abnormality, but this class mice is under the condition of normal diet, grow into 6 months when big with littermate wild-type mice and compare its body weight obviously fat partially, blood glucose increases in the blood, and sugar tolerance experiment shows that the sugared toleration of mutant mice obviously descends.Above experimental result shows that the shortage of PLRP1 gene function will significantly increase the tendency that fat and diabetes take place mice.This result will for PLRP1 in human obesity and diagnosis of diabetes and treatment, and relevant drug development aspect provides direct experimental evidence, has important use and is worth.
In the present invention, the example of non-human mammal comprises (but being not limited to): mice, rat, rabbit, monkey etc. more preferably are rat and mice.
As used herein, term " obesity/diabetes " refers to obesity and/or diabetes.
As used herein, term " PLRP1 gene " refer to encode pancreatic lipase associated protein 1 (Pancreaticlipase related proteins 1, gene PLRP1).For example, the protein sequence of people's pancreatic lipase associated protein 1 number is: ENSP00000351695, its concrete sequence is shown in SEQ ID NO:1.The protein sequence of mice pancreatic lipase associated protein 1 number is: ENSMUSP00000045465, its concrete sequence is shown in SEQID NO:2.Should be understood that this term also comprises the variant form of various naturally occurring PLRP1 genes.Representational example comprises: because of the degeneracy of the codon proteic nucleotide sequence of the PLRP1 identical of encoding with wild type, and the nucleotide sequence of the proteic conservative variation of encoding wild type PLRP1 polypeptide.In addition, during for other mammals outside the mice, this term refers to the homologue of PLRP1 gene in this mammal.For example for the people, this term refers to people's PLRP1 (research shows that the PLRP1 of mice is the lineal congener of human PLRP1, and both homologys on aminoacid are up to 83%, the gene expression profile unanimity).
In the present invention, " PLRP1 proteic conservative variation polypeptide " refers to compare with the proteic aminoacid sequence of wild type PLRP1, has 10 at the most, preferably at the most 8, more preferably at the most 5,3 aminoacid is replaced by similar performance or close aminoacid and is formed polypeptide at the most best.These conservative variation polypeptide preferably carry out the aminoacid replacement according to table 1 and produce.
Table 1
| Initial residue | Representational replacement | The preferred replacement |
| Ala(A) | Val；Leu；Ile | Val |
| Arg(R) | Lys；Gln；Asn | Lys |
| Asn(N) | Gln；His；Lys；Arg | Gln |
| Asp(D) | Glu | Glu |
| Cys(C) | Ser | Ser |
| Gln(Q) | Asn | Asn |
| Glu(E) | Asp | Asp |
| Gly(G) | Pro；Ala | Ala |
| His(H) | Asn；Gln；Lys；Arg | Arg |
| Ile(I) | Leu；Val；Met；Ala；Phe | Leu |
| Leu(L) | Ile；Val；Met；Ala；Phe | Ile |
| Lys(K) | Arg；Gln；Asn | Arg |
| Met(M) | Leu；Phe；Ile | Leu |
| Phe(F) | Leu；Val；Ile；Ala；Tyr | Leu |
| Pro(P) | Ala | Ala |
| Ser(S) | Thr | Thr |
| Thr(T) | Ser | Ser |
| Trp(W) | Tyr；Phe | Tyr |
| Tyr(Y) | Trp；Phe；Thr；Ser | Phe |
| Val(V) | Ile；Leu；Met；Phe；Ala | Leu |
As used herein, term " PLRP1 gene inactivation " comprises one or two PLRP1 gene by the situation of inactivation, promptly comprises PLRP1 genetic heterozygosis ground and isozygotys the ground inactivation.For example, the mice of PLRP1 gene inactivation can be the heterozygosis or the mice of isozygotying.
In the present invention, but gene knockout or change exogenous gene over to and make methods such as PLRP1 gene inactivation prepare the non-human mammal (as mice) of PLRP1 gene inactivation.In the art, by gene knockout or to change the technology that exogenous gene makes the target gene inactivation over to be known, these routine techniquess all can be used for the present invention.
In a preference of the present invention, the inactivation of PLRP1 gene is realized by gene knockout.
In another preference of the present invention, the inactivation of PLRP1 gene is realized by inserting exogenous gene in the PLRP1 gene.
With isozygotying of obtaining of the inventive method or the mice of heterozygosis can educate, grow normal.The PLRP1 gene inactivation of inactivation can Mendel's rule entail the offspring mice.
In a preference, the invention provides the mice of isozygotying of a kind of somatic cell and germinal cell absence PLRP1 gene, this mice can be used as obesity/diabetic mice model, be used for the research of lipid metabolism and carbohydrate metabolism, and can be used to screen the medicine that prevents and/or treats obesity and/or diabetes, especially prevent and/or treat the medicine of obesity and/or diabetes by regulation and control PLRP1 gene.
Major advantage of the present invention is:
(a) inheritance stability of obesity of the present invention/diabetic mice model, phenotype are stablized.
(b), grow normal with isozygotying of obtaining of the inventive method or the mice of heterozygosis can educate.The PLRP1 gene inactivation of inactivation can Mendel's rule entail the offspring mice.
Below in conjunction with specific embodiment, further set forth the present invention.Should be understood that these embodiment only to be used to the present invention is described and be not used in and limit the scope of the invention.The experimental technique of unreceipted actual conditions in the following example, usually according to people such as normal condition such as Sambrook, molecular cloning: laboratory manual (New York:Cold Spring Harbor Laboratory Press, 1989) condition described in, or the condition of advising according to manufacturer.
The structure of targeting vector and the preparation of gene knockout mice and evaluation
The target practice strategy as shown in Figure 1.
Targeting vector design as figure, the length of homologous recombination arm is 5.7kb, the result of homologous recombination has replaced 2-8 exon of mice PLRP1 gene with screening-gene Neo.The DNA that makes up homology arm is obtained by PCR, derives from 129 mouse ES cells DNA.Vector construction and ES cell targeting method are according to the document of delivering.
In brief, with the ES cell DNA is template, by pcr amplification homology left arm (1.6k) and right arm (4.1k), left arm is used behind XbaI and the XhoI double digestion and the P12-neo carrier (US6 of the routine of same double digestion, 528,314) connect conversion, carrier and right arm after making up with KpnI and BamHI double digestion then, connection transforms the structure of promptly finishing targeting vector again.With electrotransfection ES cell after the targeting vector linearisation, obtain the cell clone of anti-G418.Extract the DNA of each cell clone, use the generation of the method evaluation homologous recombination of PCR, obtain the needed positive ES cell clone that PLRP1 gene delection has taken place.The positive ES cell clone of cultivating is expelled in the mice segmentation cavity, again with embryo transfer in the pseudo-fetus Mouse Uterus of synchronization of estrus, continue to grow, obtain gomphosis mouse.Gomphosis mouse carries out heredity to it and breeds after obtaining, and obtains heterozygote mice and the homozygote mice of PLRP1 at last.Can utilize mice tail end tissue extraction DNA, and utilize PCR to distinguish the mice of different genotype.The PCR primer of distinguishing wild type, heterozygote and homozygote mice is as follows respectively:
P1：5’-GGGCCCCACCATGCTTGCTCT-3’(SEQ?ID?NO：3)
P2：5’-TCGGCAGGAGCAAGGTGAGATGACAGGAG-3’(SEQ?ID?NO：4)
P3：5’-CCACCGGGACCTTTTTATGCTC-3’(SEQ?ID?NO：5)
In PCR when reaction, add above-mentioned 3 kinds of primers simultaneously, corresponding genotype as shown in Figure 2, wherein the homozygote mice has the reaction band of 1.5kb, wild-type mice has the band of 1.0kb, heterozygote has above-mentioned two simultaneously.
Adopt Western Blot method to gene knockout mice PLRP1 proteic removal identify that further method is as follows:
Pancreas extracting albumen is transferred on the nitrocellulose filter behind SDS polyacrylate hydrogel electrophoresis.WesternBlot carries out according to conventional methods.Antibody is the anti-mice PLRP1 of the rabbit multi-resistance with the conventional method preparation.
The result shows, detected less than PLRP1 is proteic in the PLRP1 gene knockout homozygote mice (KO) to have (Fig. 3).
PLRP1 gene knockout mice fat content changes research
To wild-type mice and PLRP1 gene knockout mice, feed with normal diet, when the mice age is 6.5 months, measure fat content with desk-top magnetic nuclear resonance analyzer Bruker minispec (available from Bruker company).
The result as shown in Figure 4, the fat content of gene knockout mice (KO) is greater than wild type (WT).This shows that the inactivation of PLRP1 can be causeed fat.
Embodiment 3
The research of PLRP1 gene knockout mice change of blood sugar
To wild-type mice and PLRP1 gene knockout mice, feed with normal diet, when the mice age is 6.5 months, measure blood glucose value with the biochemistry analyzer of Hitachi's board.
The result as shown in Figure 5, the blood glucose value of gene knockout mice (KO) is greater than wild type (WT).This shows that the inactivation of PLRP1 can cause hyperglycemia.
The homology of human PLRP1 protein sequence and mice PLRP1 protein sequence relatively
With the blastp software (http://www.ncbi.nlm.nih.gov/BLAST/) of NCBI, PLRP1 (SEQ ID NO:2 and Fig. 7) of mice and people's PLRP1 (SEQ ID NO:1 and Fig. 6) are carried out sequence alignment.
Comparison result as shown in Figure 8, both homologys on aminoacid sequence are up to 83%.
The chemical compound of screening preventing/treating obesity or hyperglycemia
In matched group,, feed with normal diet to wild-type mice and PLRP1 gene knockout mice.In the test group, to the PLRP1 gene knockout mice, feed, and regularly add candidate's test substances (plant extract or synthetic chemical compound) with normal diet.When the mice age is 6 months, measure fat content with embodiment 4 same procedure, measure blood glucose value with embodiment 5 same procedure.
If fat content significantly is lower than the PLRP1 gene knockout mice in the matched group in the test group, then point out this test compounds that the effect that reduces fat content is arranged, can be used as the drug candidate of preventing/treating obesity.
If blood glucose value content significantly is lower than the PLRP1 gene knockout mice in the matched group in the test group, the effect of then pointing out this test compounds that blood sugar lowering is arranged can be used as the drug candidate of preventing/treating hyperglycemia.
Discuss
After the Human Genome Project was finished, the center of gravity of life science expanded to the functional selection of whole animal and application clinically thereof from molecule and gene level.Wherein model animal comprises that fruit bat, nematicide, Brachydanio rerio, mice etc. are topmost object of study.Mice genome and the human homology that has more than 90%, mice is very similar to the mankind at fetal development, histoorgan and function, the conduction of cell internal information and biochemical metabolism path etc., through the mouse model that genetic engineering is modified, be that gene function, human diseases mechanism and new drug research are developed most important model organism.As the model of human diseases, mouse model has been contained the animal model of multiple human diseasess such as cardiovascular system obstacle such as comprising diabetes, obesity, atrial fibrillation.
Utilize the function of mouse model research unknown gene, be by gene knockout, gene mutation or transgenic, produce mice through genetic modification, by the analysis that the animal phenotype is changed and the mutation analysis of Physiological and Biochemical Metabolism process, resolve the function of gene at whole animal, cell and molecular level, thus with related gene as pharmaceutically-active target spot, be used for carrying out the screening of newtype drug.
For PLRP1, there are some researches show, need a large amount of PLRP1 albumen and various substrate, could find to have the esterase active of trace, this active bile salt and the colipase of relying on, that interesting is active suitable (the Crenon I with lipase of combination of PLRP1 and colipase, Foglizzo E, Kerfelec B, Verine A, Pignol D, Hermoso J, Bonicel J, Chapus C.Pancreatic lipase-related protein typeI:a specialized lipase or aninact ive enzyme.Protein Eng.1998Feb; 11 (2): 135-42.).
In fact, PLRP1 is in structure and aminoacid is formed and PL is closely similar, structure and sequence according to PL, only the V179 of PLRP1 and A181 need be mutated into activity (the Crenon I that A179 and P181 just can significantly rebuild the PLRP1 esterase, Jayne S, Kerfelec B, Hermoso J, Pignol D, ChapusC.Pancreatic lipase-related protein type1:a double mutation restoresa significant lipase activity.Biochem Biophys Res Commun.1998May19; 246 (2): 513-7; Bezzine S, Roussel A, de Caro J, Gastinel L, de CaroA, Carriere F, Leydier S, Verger R, Cambillau C.An inactive pancreaticlipase-related protein is activated into a triglyceride-lipase bymutagenesis based on the3-D structure.Chem Phys Lipids.1998Jun; 93 (1-2): 103-14.).
According to above-mentioned result of study, the inventor thinks that PLRP1 may be the active albumen of regulating of of PL, suppresses the activity of PL in conjunction with colipase by competition.What some were external studies show that, the PL albumen of inactivation suppresses the activity of PL consumingly by the competition colipase.
In addition, PLRP1 albumen mainly is expressed in the pancreas, in islets of langerhans, express higher, different with PL, it promptly expresses (Yang Y the period of embryo, Sanchez D, Figarella C, Lowe ME.Discoordinateexpression of pancreatic lipase and two related proteins in the humanfetal pancreas.Pediatr Res.2000Feb; 47 (2): 184-8.).
There is polymorphism in the PLRP1 gene in the crowd, but also not with its evidence (Cao H, Hegele RA DNA polymorphisms of lipase related genes.J HumGenet.2003 with human metabolic disease contact; 48 (8): 443-6.Epub2003Aug1).
PLRP1 is subjected to the adjusting of cortisone, the expression of cortisone downward modulation PLRP1, and but expression (the Kullman J of rise PLRP2, Gisi C, Lowe ME.Dexamethasone-regulated expression ofpancreatic lipase and two related proteins in AR42J cells.Am J Physiol.1996May; 270 (5Pt1): G746-51), and leptin (leptin) also can be reduced the expression of PL and PLRP1, raise expression (the Elinson N of PLRP2, Amichay D, Birk RZ.Leptin directlyregulates exocrine pancreas lipase and two related proteins in the rat.Br J Nutr.2006Oct; 96 (4): 691-6.), the meaning of PLRP1 albumen aspect the hormonal regulation metabolism is also indeterminate.
The inventor utilizes the dna homology recombinant technique, has prepared the mice of rejecting the gene of PLRP1 first.PLRP1-/-mice can normally be born, grow and maturation, and the energy procreate.The diet of homozygote mice contrasts with wild type with motion and compares Non Apparent Abnormality, but this class mice is under the condition of normal diet, grow into 6 months when big with littermate wild-type mice and compare its body weight obviously fat partially, blood glucose increases in the blood, and sugar tolerance experiment shows that the sugared toleration of mutant mice obviously descends.Above experimental result shows that the shortage of PLRP1 gene function will significantly increase the tendency that fat and diabetes take place mice.This result will for PLRP1 in human obesity and diagnosis of diabetes and treatment, and relevant drug development aspect provides direct experimental evidence, has important use and is worth.
All quote in this application as a reference at all documents that the present invention mentions, just quoted as a reference separately as each piece document.Should be understood that in addition those skilled in the art can make various changes or modifications the present invention after having read above-mentioned teachings of the present invention, these equivalent form of values fall within the application's appended claims institute restricted portion equally.
Sequence table
＜110〉Shanghai Nanfangmoshi Biological Sci-Tech Dev Co., Ltd.
Model organism research center, south, Shanghai
＜120〉purposes of pancreatic lipase associated protein 1
<130>071989
<160>5
<170>PatentIn?version3.4
<210>1
<211>467
<212>PRT
＜213〉people (homo sapiens)
<400>1
<210>2
<211>473
<212>PRT
＜213〉mice (Mus musculus)
<400>2
<210>3
<211>21
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
＜223〉primer
<400>3
<210>4
<211>29
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
＜223〉primer
<400>4
<210>5
<211>22
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
＜223〉primer
<400>5
Claims (6)
1. a method that produces the non-human mammal embryonic stem cell of obesity/diabetes is characterized in that, comprises step:
(a) by rejecting the PLRP1 gene or insert exogenous gene from genome in the PLRP1 gene, produce the embryonic stem cell of the non-human mammal of PLRP1 gene inactivation, wherein PLRP1 is a pancreatic lipase associated protein 1.
2. the method for claim 1 is characterized in that, described step (a) is to cause the PLRP1 inactivation by the exon of replacing the PLRP1 gene with the Neo gene.
3. the method for claim 1 is characterized in that, described non-human mammal is mice, rat, rabbit, monkey.
4. the method for claim 1 is characterized in that, two PLRP1 genes in the genome of the embryonic stem cell of non-human mammal wherein are all owing to rejecting or inserting exogenous gene and inactivation.
5. the purposes with the embryonic stem cell of the non-human mammal of the described method acquisition of claim 1 is characterized in that, is used to prepare obesity/diabetes model; Or be used to screen and prevent and/or treat fat medicine; And/or be used to screen the medicine of prevention and/or diabetes.
6. purposes as claimed in claim 5 is characterized in that, described non-human mammal is mice or rat.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2007100435817A CN101342372B (en) | 2007-07-09 | 2007-07-09 | Use of pancreatic lipase associated protein 1 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2007100435817A CN101342372B (en) | 2007-07-09 | 2007-07-09 | Use of pancreatic lipase associated protein 1 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101342372A CN101342372A (en) | 2009-01-14 |
| CN101342372B true CN101342372B (en) | 2011-06-29 |
Family
ID=40244563
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2007100435817A Active CN101342372B (en) | 2007-07-09 | 2007-07-09 | Use of pancreatic lipase associated protein 1 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101342372B (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1580266A (en) * | 2003-08-13 | 2005-02-16 | 上海南方模式生物科技发展有限公司 | Method for extablishing polymorphism adenoid tumour mouse model |
| CN1795929A (en) * | 2004-12-30 | 2006-07-05 | 上海南方模式生物科技发展有限公司 | Method for generating model of chmice in male sterile |
| CN1289662C (en) * | 2003-08-08 | 2006-12-13 | 上海南方模式生物科技发展有限公司 | Method for efficiently establishing pure line gene knock-out mouse model |
-
2007
- 2007-07-09 CN CN2007100435817A patent/CN101342372B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1289662C (en) * | 2003-08-08 | 2006-12-13 | 上海南方模式生物科技发展有限公司 | Method for efficiently establishing pure line gene knock-out mouse model |
| CN1580266A (en) * | 2003-08-13 | 2005-02-16 | 上海南方模式生物科技发展有限公司 | Method for extablishing polymorphism adenoid tumour mouse model |
| CN1795929A (en) * | 2004-12-30 | 2006-07-05 | 上海南方模式生物科技发展有限公司 | Method for generating model of chmice in male sterile |
Also Published As
| Publication number | Publication date |
|---|---|
| CN101342372A (en) | 2009-01-14 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Greer et al. | Hoxb8 is required for normal grooming behavior in mice | |
| CN100381570C (en) | Synovial membrane cell protein | |
| DE10294485T5 (en) | Proteins homologous with Mnk kinase, which are involved in the regulation of energy homeostasis and organelle metabolism | |
| US6734337B2 (en) | Acetyl-coenzyme A carboxylase 2 as a target in the regulation of fat burning, fat accumulation, energy homeostasis and insulin action | |
| CN110894508B (en) | Application of gene Adra1a for regulating browning of white fat | |
| Lu et al. | The mammalian retinal degeneration B2 gene is not required for photoreceptor function and survival | |
| CN102482337A (en) | Muteins of the pyrroline-5-carboxylate reductase 1 | |
| Fortier et al. | Human hormone-sensitive lipase (HSL): expression in white fat corrects the white adipose phenotype of HSL-deficient mice | |
| Wu | Molecular phyloecology suggests a trophic shift concurrent with the evolution of the first birds | |
| CN101342372B (en) | Use of pancreatic lipase associated protein 1 | |
| US20110212081A1 (en) | Krüppel-like factors and fat regulation | |
| Kiemnec-Tyburczy et al. | Evolutionary shifts in courtship pheromone composition revealed by EST analysis of plethodontid salamander mental glands | |
| US6740793B2 (en) | Transgenic animal having a disrupted PDE7A gene and uses thereof | |
| US20040204338A1 (en) | Acetyl-coenzyme a carboxylase 2 as a target in the regulation of fat burning, fat accumulation, energy homeostasis and insulin action | |
| EP1605965B1 (en) | Use of saposin-related proteins for preventing and treating obesity, diabetes and/or metabolic syndrome | |
| CN113862305A (en) | Construction method of ATP7B gene knockout mouse model | |
| US20220007621A1 (en) | Animal model of mucopolysaccharidoses type iva | |
| US6602698B2 (en) | Human kinase proteins and polynucleotides encoding the same | |
| Gu et al. | Targeted over-expression of PPARγ in pig skeletal muscle improves oxidative fiber formation and intramuscular fat deposition | |
| Fukamizu | Transgenic animals in endocrinological investigation | |
| Ji AiBing et al. | Breeding of leptin receptor gene knockout mice and genotype identification of the offspring. | |
| Shintani et al. | Overexpression of PACAP in the pancreas failed to rescue early postnatal mortality in PACAP-null mice | |
| WO2022221299A1 (en) | Insect products and methods of producing same | |
| WO2005051408A1 (en) | Method for preventing and treating diabetes using dg119 | |
| CN115349015A (en) | Agents and methods for regulating sleep |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20200106 Address after: 201203 Shanghai Jinke Road, Zhangjiang High Tech Park of Pudong New Area No. 3577 Building No. 2 Co-patentee after: SHANGHAI LAB. ANIMAL RESEARCH CENTER Patentee after: SHANGHAI RESEARCH CENTER FOR MODEL ORGANISMS Address before: 201203 Shanghai science and Technology Industrial Zone Zhangjiang Cailun Road No. 88 2 4 floor Co-patentee before: SHANGHAI RESEARCH CENTER FOR MODEL ORGANISMS Patentee before: SHANGHAI BIOMODEL ORGANISM SCIENCE & TECHNOLOGY DEVELOPMENT Co.,Ltd. |
|
| TR01 | Transfer of patent right | ||
| CP03 | Change of name, title or address |
Address after: 201318 floor 2-4, building 2, No. 178, Banxia Road, Pudong New Area, Shanghai Co-patentee after: SHANGHAI RESEARCH CENTER FOR MODEL ORGANISMS Patentee after: SHANGHAI MODEL ORGANISMS CENTER, Inc. Address before: 201203 Shanghai science and Technology Industrial Zone Zhangjiang Cailun Road No. 88 2 4 floor Co-patentee before: SHANGHAI RESEARCH CENTER FOR MODEL ORGANISMS Patentee before: SHANGHAI BIOMODEL ORGANISM SCIENCE & TECHNOLOGY DEVELOPMENT Co.,Ltd. |
|
| CP03 | Change of name, title or address | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20200331 Address after: 201318 floor 2-4, building 2, No. 178, Banxia Road, Pudong New Area, Shanghai Co-patentee after: SHANGHAI LAB. ANIMAL RESEARCH CENTER Patentee after: SHANGHAI MODEL ORGANISMS CENTER, Inc. Address before: 201318 floor 2-4, building 2, No. 178, Banxia Road, Pudong New Area, Shanghai Co-patentee before: SHANGHAI RESEARCH CENTER FOR MODEL ORGANISMS Patentee before: SHANGHAI MODEL ORGANISMS CENTER, Inc. |
|
| TR01 | Transfer of patent right |