CN101812525A - Method for building Alzheimer's disease (AD) morbidity-associated gene network path analyzing model - Google Patents
Method for building Alzheimer's disease (AD) morbidity-associated gene network path analyzing model Download PDFInfo
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Abstract
The invention discloses a method for building an Alzheimer's disease (AD) morbidity-associated gene network path analyzing model, which comprises the following steps of: collecting the known AD morbidity-associated genes by using a bioinformatics method; performing analysis of genetic genomics by means of gene expression data and gene-type data of B*D gene recombinant inbred mice to screen out upstream regulating loci with statistical significance; performing verification of cis-eQTL authenticity and candidate genes screening verification; and determining the synergic or regulatory relation of the AD morbidity-associated genes to build the gene expression regulating network path of the AD morbidity-associated genes. The method is simple and convenient and easy to operate; and the AD morbidity-associated gene network path analyzing model can be effectively built by the method.
Description
Technical field:
The present invention relates to a kind of method of the AD of structure pathogenesis related genes network path.
Background technology:
Alzheimer's disease (Alzheimer disease, AD) be a kind of modal central nervous system degenerative disease, raising along with people's living standard, human longevity constantly prolongs, aging degree is aggravated gradually, diseases such as AD have influenced the elderly's health and life more and more significantly, have become one of major disease of modern society's serious harm human health.
The main pathology of AD is characterized as cerebral atrophy, form in the cerebral tissue senile plaque (senileplaque, SP), neurofibrillary tangles (neurofilament tangles, NFTs) and cerebrovascular throw out etc.The Neuropathological Study result of AD patient's cerebral tissue shows, there is the forfeiture of cholinergic widely or noradrenergic system moiety in positions such as pallium, hippocampus, basal forebrain, nucleus ceruleus, and wherein hippocampus is the major lesions functional zone of AD.Modern medicine study shows that first symptom of AD shows as and the closely-related lethe of hippocampus function; Simultaneously people have found also that the early stage pathologic of AD changes and occur in hippocampus CA1 district or occur in hippocampus simultaneously and adjacent with the hippocampus cortex of frontal lobe (entorhinal cortex) that sends projection fibre to hippocampus, along with the development of the state of an illness, pathological change just is extended to basal forebrain position nuclear groups such as other cortex and amygdala gradually.
The cause of disease of AD is very complicated, but increases age and inherited genetic factors is known together by everybody.Because the heterogeneity in the AD heredity makes all unusual difficulties of the prevention of AD and diagnosis.Usually AD is divided into 3 classes clinically: familial is early sent out, familial is sent out and sent out sporadic evening evening.Existing study of pathogenesis to AD mainly concentrates on carries out single-gene or single proteic further investigation to AD relevant oligogene and functional protein thereof, or the characteristic pathological product NFTs of AD and the forming process of SP are done careful research.At present existing many scholars have proposed many hypothesis, (Nussbaum RL and Ellis CE.Alzheimer ' sdisease and Parkinson ' s disease.N Engl J Med such as, Protein tau excessive phosphorylation too much, peroxidation, inflammatory factor, metal and the disorder of intracellular Ca2+ stable state, inherited genetic factors as A β product, 2003,348 (14): 1356-1364).The proposition of these hypothesis is to deeply being familiar with the very big promoter action that truly has of AD pathogenesis, the effective methods of treatment of exploration etc.But owing to lack the research of whole and system level, AD morbidity Molecular Study also fails to make a breakthrough, and does not also have ideal specificity early diagnosis biology sign so far, and the control of AD remains a global difficult problem.Therefore, we think that research will carry out on level in integral body or the system, and the idiotype network regulatory mechanism of AD morbidity is analyzed in the especially systematic study on the genome aspect, and the morbidity of people being understood in depth AD will be very helpful.
Biochip technology previously mainly detects the difference of thousands of genetic expression, only can analyze on the aspect of genetic expression, so as to inferring genetic expression and a series of biological actions that product produced thereof; And people often utilize biochip technology that the analysis of hanging down quantity of information, poor efficiency is analyzed in the regulation and control of single or a few genetic expression isolatedly, so can't fully understand the network regulation of genomic genetic expression and related gene expression.Calendar year 2001, Jansen and Nap propose and can regard the mRNA expression amount of each gene in the full genome as quantitative character, it is carried out quantitative character location (quantitative trait loci, QTL) analyze, promptly in conjunction with the genetic expression quantitative trait locus positioning analysis of qtl analysis and gene expression analysis.2002, people such as Brem took the lead in this research means successful Application in zymic research; 2003, people such as Schadt used this method and seek the Disease-causing gene path in the F2 mouse.The analysis integrated method of using genomics, statistical genetics and information biology of eQTL, can be used for seeking the upstream regulatory site of certain genetic expression of control in the genome, excavation be subjected to this generegulation downstream gene and with other genes of this synergistic effect of gene, set up the gene expression regulation network, the hereditary basis of research changes in gene expression on genetic expression and two levels of expression regulation, and then the molecule mechanism and the regulated and control network of announcement complex character.Analyze by eQTL, we both can understand the difference of most of genetic expressions by biochip technology, can find the upstream gene site of these gene expression differences of control simultaneously again by means of the QTL location technology.As for genetic expression quantitative trait locus location technology is applied in the research of diseases such as AD, also there is not relevant report at present both at home and abroad.
Summary of the invention:
The object of the present invention is to provide the establishment method of the relevant idiotype network path analysis model of a kind of and AD morbidity.
Technical solution of the present invention is:
The establishment method of the idiotype network path analysis model that a kind of and AD morbidity is relevant is characterized in that: comprise the following steps:
By bioinformatics method, collect known AD genes involved; By B * D gene recombination inbred mouse (Recombinant Inbred, RI) genetic expression (expression) data and genotype (genotype) data, carry out the analysis that the gene group is learned, filter out upstream regulatory site with statistical significance; Carry out cis-eQTL authenticity verification, candidate gene screening verification; Determine the collaborative or regulation relationship between the AD pathogenesis related genes, make up the gene expression regulation network path of AD genes involved.
Collect known AD pathogenesis related genes, main by retrieval Pubmed (http://www.ncbi.nlm.nih.gov/sites/entrez/), web of Science (http://www.isiknowledge.com/), Chilibot databases such as (http://www.chilibot.net/), consult and analyze pertinent literature, collect the AD candidate gene, and select wherein and analyze for most scholar's approvals or through the candidate gene of lot of experiment validation.
Described gene expression data and genotype data by B * D gene recombination inbred mouse carried out correlation analysis, and the means of wherein obtaining gene expression data comprise the method for biochip technology, RT-PCR and the method for ABA in situ hybridization; Use Mit (Microsatellites), SNP (Single Nucleotide Polymorphism) equimolecular genetic marker that B * D gene recombination inbred mouse is carried out full genome scanning, obtain the genotype data.
The described analysis of carrying out gene group, filter out upstream regulatory site with statistical significance, be that gene recombination inbred mouse to B * D carries out the localized preliminary analysis of genetic expression quantitative trait locus, filter out upstream regulatory site with statistical significance.
The described cis-eQTL authenticity verification that carries out comprises the following steps:
(1) gene expression amount is carried out the QTL The Location, according to eQTL position and gene self position distance, at a distance of the 5Mb scope with the interior cis-acting expression quantitative trait locus that is considered to, otherwise be trans-acting genetic expression quantitative trait locus;
(2) investigate the gene expression amount of two parent B6 and D2, reject the gene of differential expression not statistically significant between two parents;
(3) investigate the polymorphism of probe target sequence, if there is pleomorphism site, and this site is to cause the gene of higher LRS then to reject;
(4) carry out the allele-specific expression analysis, then can think true property cis-eQTL if the differential expression between two allelotrope B6 and D2 has statistical significance.
The screening of described candidate gene and checking, step is as follows:
(1) cis-eQTL is behind authenticity verification, and its candidate gene is this gene self;
(2) candidate gene of trans-eQTL can screen through following method:
I. be positioned at the gene in trans-eQTL interval, if certainly as the cis regulatory gene, this cis regulatory gene is candidate gene probably;
Ii. by increasing the density of local genome scanning, dwindle 95% credibility interval of upstream regulatory site;
Iv. filter out the gene relevant by bioinformatics method with growth, structure and the function of hippocampus;
V. analyze gene expression difference between two parent B6 and D2, filter out the gene that gene expression amount has notable difference;
Vi. by means of Celera, Perlegen and Tennessee State university snp database, filter out the gene that between B6 and D2, has functional SNP;
Vii. use the luciferase technology in the influence of the difference of external checking exon to genetic expression;
Viii. construction expression plasmid, the intergenic adjusting relation in external checking upstream and downstream;
Ix. use gene knockout or gene weakening technology that the gene that part may exist upstream and downstream to regulate relation is verified, regulate in the path in the idiotype network of having set up, use the related gene knock-out mice, verify by the variation of comparing the upstream and downstream genetic expression in normal mouse and this gene pathway of knock out mice whether gene pathway exists; Or the related gene in the application RNA perturbation technique suppressor gene network, to observe the variation of its upstream and downstream genetic expression.
The gene expression regulation network path of described structure AD genes involved is by the software and the website that are specifically designed to the idiotype network analysis upstream candidate gene and its downstream regulated gene to be analyzed, and sets up the network adjustment path between the genes involved; There are many genes may not exist direct upstream and downstream to regulate relation, but they are closely related on function, by gene expression data is carried out the genetics correlation analysis, find out and the gene relevant with known AD morbidity between have the gene of height correlation, determine as yet report not may with other relevant gene of AD morbidity.
The inventive method is easy, and is easy to operate, can effectively set up and the relevant idiotype network path analysis model of AD morbidity.
Description of drawings:
The invention will be further described below in conjunction with drawings and Examples.
Fig. 1 is the schema of the present invention to gene group research means implementation procedure.
Embodiment:
The establishment method of the idiotype network path analysis model that a kind of and AD morbidity is relevant comprises the following steps:
By bioinformatics method, collect known AD genes involved; By B * D gene recombination inbred mouse (Recombinant Inbred, RI) genetic expression (expression) data and genotype (genotype) data, carry out the analysis that the gene group is learned, filter out upstream regulatory site with statistical significance; Carry out cis-eQTL authenticity verification, candidate gene screening verification; Determine the collaborative or regulation relationship between the AD pathogenesis related genes, make up the gene expression regulation network path of AD genes involved.
Described B * D gene recombination inbred mouse is to obtain gene recombination inbred mouse B * D RI mouse by the hybridization more than continuous 20 generations of two parent B6 and D2.
Collect known AD pathogenesis related genes, main by retrieval Pubmed (http://www.ncbi.nlm.nih.gov/sites/entrez/), web of Science (http://www.isiknowledge.com/), Chilibot databases such as (http://www.chilibot.net/), consult and analyze pertinent literature, collect the AD candidate gene, and select wherein and analyze for most scholar's approvals or through the candidate gene of lot of experiment validation.
Described gene expression data and genotype data by B * D gene recombination inbred mouse carried out correlation analysis, and the means of wherein obtaining gene expression data comprise that the method for the method of biochip technology, RT-PCR and ABA in situ hybridization obtains the gene expression data of B * D RI mouse hippocampal tissue; Use Mit (Microsatellites), SNP (Single NucleotidePolymorphism) equimolecular genetic marker that B * D gene recombination inbred mouse is carried out full genome scanning, obtain the genotype data.
The described analysis of carrying out gene group, filter out upstream regulatory site with statistical significance, be that gene recombination inbred mouse to B * D carries out the localized preliminary analysis of genetic expression quantitative trait locus, filter out upstream regulatory site with statistical significance.
The described cis-eQTL authenticity verification that carries out comprises the following steps:
(1) gene expression amount is carried out the QTL The Location, according to eQTL position and gene self position distance, at a distance of the 5Mb scope with the interior cis-acting expression quantitative trait locus that is considered to, otherwise be trans-acting genetic expression quantitative trait locus;
(2) investigate the gene expression amount of two parent C57BL/6J (B6) and DBA/2J (D2), reject the gene of differential expression not statistically significant between two parents;
(3) by means of Celera, Perlegen and Tennessee State university snp database, the polymorphism of analysis probe target sequence, if there is pleomorphism site, and this site is to cause the gene of higher LRS then to reject;
(4) carry out the allele-specific expression analysis, then can think true property cis-eQTL if the differential expression between two allelotrope B6 and D2 has statistical significance.
The screening of described candidate gene and checking, step is as follows:
(1) cis-eQTL is behind authenticity verification, and its candidate gene is this gene self;
(2) candidate gene of trans-eQTL can screen through following method:
I. be positioned at the gene in trans-eQTL interval, if certainly as the cis regulatory gene, this cis regulatory gene is candidate gene probably;
Ii. by increasing the density of local genome scanning, dwindle 95% credibility interval of upstream regulatory site;
Iv. filter out the gene relevant by bioinformatics method with growth, structure and the function of hippocampus;
V. analyze gene expression difference between two parent B6 and D2, filter out the gene that gene expression amount has notable difference;
Vi. by means of Celera, Perlegen and Tennessee State university snp database, filter out the gene that between B6 and D2, has functional SNP;
Vii. use the luciferase technology in the influence of the difference of external checking exon to genetic expression;
Viii. construction expression plasmid, the intergenic adjusting relation in external checking upstream and downstream;
Ix. use gene knockout or gene weakening technology that the gene that part may exist upstream and downstream to regulate relation is verified, regulate in the path in the idiotype network of having set up, use the related gene knock-out mice, verify by the variation of comparing the upstream and downstream genetic expression in normal mouse and this gene pathway of knock out mice whether gene pathway exists; Or the related gene in the application RNA perturbation technique suppressor gene network, to observe the variation of its upstream and downstream genetic expression.
The gene expression regulation network path of described structure AD genes involved, be to pass through WebGestalt, Ingenuity etc. are specifically designed to the software and the website of idiotype network analysis upstream candidate gene and its downstream regulated gene are analyzed, and set up the network adjustment path between the genes involved; There are many genes may not exist direct upstream and downstream to regulate relation, but they are closely related on function, by gene expression data is carried out the genetics correlation analysis, find out and the gene relevant with known AD morbidity between have the gene of height correlation, determine as yet report not may with other relevant gene of AD morbidity.
Claims (6)
1. the establishment method of an idiotype network path analysis model relevant with the AD morbidity is characterized in that: comprise the following steps:
By bioinformatics method, collect known AD genes involved; By the gene expression data and the genotype data of B * D gene recombination inbred mouse, carry out the analysis that the gene group is learned, filter out upstream regulatory site with statistical significance; Carry out cis-eQTL authenticity verification, candidate gene screening verification; Determine the collaborative or regulation relationship between the AD pathogenesis related genes, make up the gene expression regulation network path of AD genes involved.
2. the establishment method of the idiotype network path analysis model that according to claim 1 and AD morbidity is relevant, it is characterized in that: described gene expression data and genotype data by B * D gene recombination inbred mouse carried out correlation analysis, and the means of wherein obtaining gene expression data comprise the method for biochip technology, RT-PCR and the method for ABA in situ hybridization; Use Mit, SNP molecular genetic sign that B * D gene recombination inbred mouse is carried out full genome scanning, obtain the genotype data.
3. the establishment method of the idiotype network path analysis model that according to claim 1 and 2 and AD morbidity is relevant, it is characterized in that: the described analysis of carrying out gene group, filter out upstream regulatory site with statistical significance, be that gene recombination inbred mouse to B * D carries out the localized preliminary analysis of genetic expression quantitative trait locus, filter out upstream regulatory site with statistical significance.
4. the establishment method of the idiotype network path analysis model that according to claim 1 and 2 and AD morbidity is relevant, it is characterized in that: the described cis-eQTL of carrying out authenticity verification comprises the following steps:
(1) gene expression amount is carried out the QTL The Location, according to eQTL position and gene self position distance, at a distance of the 5Mb scope with the interior cis-acting expression quantitative trait locus that is considered to, otherwise be trans-acting genetic expression quantitative trait locus;
(2) investigate the gene expression amount of two parent B6 and D2, reject the gene of differential expression not statistically significant between two parents;
(3) investigate the polymorphism of probe target sequence, if there is pleomorphism site, and this site is to cause the gene of higher LRS then to reject;
(4) carry out the allele-specific expression analysis, then can think true property cis-eQTL if the differential expression between two allelotrope B6 and D2 has statistical significance.
5. the establishment method of the idiotype network path analysis model that according to claim 1 and 2 and AD morbidity is relevant is characterized in that: the screening of described candidate gene and checking, and step is as follows:
(1) cis-eQTL is behind authenticity verification, and its candidate gene is this gene self;
(2) candidate gene of trans-eQTL can screen through following method:
I. be positioned at the gene in trans-eQTL interval, if certainly as the cis regulatory gene, this cis regulatory gene is candidate gene probably;
Ii. by increasing the density of local genome scanning, dwindle 95% credibility interval of upstream regulatory site;
Iv. filter out the gene relevant by bioinformatics method with growth, structure and the function of hippocampus;
V. analyze gene expression difference between two parent B6 and D2, filter out the gene that gene expression amount has notable difference;
Vi. by means of Celera, Perlegen and Tennessee State university snp database, filter out the gene that between B6 and D2, has functional SNP;
Vii. use the luciferase technology in the influence of the difference of external checking exon to genetic expression;
Viii. construction expression plasmid, the intergenic adjusting relation in external checking upstream and downstream;
Ix. use gene knockout or gene weakening technology that the gene that part may exist upstream and downstream to regulate relation is verified, regulate in the path in the idiotype network of having set up, use the related gene knock-out mice, verify by the variation of comparing the upstream and downstream genetic expression in normal mouse and this gene pathway of knock out mice whether gene pathway exists; Or the related gene in the application RNA perturbation technique suppressor gene network, to observe the variation of its upstream and downstream genetic expression.
6. the establishment method of the idiotype network path analysis model that according to claim 1 and 2 and AD morbidity is relevant, it is characterized in that: the gene expression regulation network path of described structure AD genes involved, be upstream candidate gene and its downstream regulated gene to be analyzed, set up the network adjustment path between the genes involved by the software and the website that are specifically designed to the idiotype network analysis; There are many genes may not exist direct upstream and downstream to regulate relation, but they are closely related on function, by gene expression data is carried out the genetics correlation analysis, find out and the gene relevant with known AD morbidity between have the gene of height correlation, determine as yet report not may with other relevant gene of AD morbidity.
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| CN104361263A (en) * | 2014-10-10 | 2015-02-18 | 北京林业大学 | Forest tree breeding method and system based on gene and protein regulatory network |
| CN105063186A (en) * | 2011-11-10 | 2015-11-18 | 霍夫曼-拉罗奇有限公司 | Methods for treating, diagnosing and monitoring Alzheimer's disease |
| CN109003673A (en) * | 2017-06-05 | 2018-12-14 | 新加坡北斗多维养生公司 | A kind of method and skin care method of the active constituent and skin care item for recommending to be suitable for individual |
| CN112802546A (en) * | 2020-12-29 | 2021-05-14 | 中国人民解放军军事科学院军事医学研究院 | Biological state characterization method, device, equipment and storage medium |
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2010
- 2010-04-09 CN CN201010150139A patent/CN101812525A/en active Pending
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105063186A (en) * | 2011-11-10 | 2015-11-18 | 霍夫曼-拉罗奇有限公司 | Methods for treating, diagnosing and monitoring Alzheimer's disease |
| CN104361263A (en) * | 2014-10-10 | 2015-02-18 | 北京林业大学 | Forest tree breeding method and system based on gene and protein regulatory network |
| CN104361263B (en) * | 2014-10-10 | 2018-03-27 | 北京林业大学 | The accurate breeding method of forest and system based on gene Yu protein regulation network |
| CN109003673A (en) * | 2017-06-05 | 2018-12-14 | 新加坡北斗多维养生公司 | A kind of method and skin care method of the active constituent and skin care item for recommending to be suitable for individual |
| CN112802546A (en) * | 2020-12-29 | 2021-05-14 | 中国人民解放军军事科学院军事医学研究院 | Biological state characterization method, device, equipment and storage medium |
| CN112802546B (en) * | 2020-12-29 | 2024-05-03 | 中国人民解放军军事科学院军事医学研究院 | Biological state characterization method, device, equipment and storage medium |
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Application publication date: 20100825 |