주름 피부 타입 유전자 다형성 마커 및 이의 용도

One aspect for achieving said purpose, the present invention refers to determining whether or not the skin wrinkles can be selected from at least one single base polymorphisms (SNP) markers 1 polymorphic marker including a marker for the diagnosis of the wrinkles of skin type single base number under public affairs substrate.

In the present invention terms, "polymorphisms (polymorphism)" RM one gene (allele) gene allele (locus) are printed in the presence of an allele of areas it is, only a single nucleotide sequence (single nucleotide polymorphism, SNP) single base along another person is combined with a load. Polymorphic marker that preferably selected population 1% or more, 10% or 20% or more indicative of the frequency of occurrence of one or more more preferably have an allele of gene.

Terms in the present invention, "gene allele (allele)" homologous chromosome gene is the same gene above left various types to bring said substrate. Gene prediction for the wherein the polymorphisms alleles is, e.g., an allele of a SNP is two factor (biallele).

Terms in the present invention, provide the means by which information recording year 1998 "rs_id" SNP from percent of the NCBI SNP that it initially all be altered independent rs a-ID a marking becoming a big. The table of the present invention is a big rs_id polymorphic marker SNP marker.

In the present invention terms, meaning that the skin type "skin type" RM to individuals having a measuring or diagnostic unit, of the present invention SNP number without measuring the skin type may include, be a preferred embodiments. The present invention according to an exemplary embodiment of the present invention the victims of the skin portion of a subject is skin wrinkles was many or corrugated skin type classification. Single base polymorphic marker of the present invention can be accurate for measuring skin type accessories to, information about a change in the skin type active ingredient be skin in contact with the ceramic.

Table 3 and table 4 preferably single base said polymorphic marker on the display is at least one selected among single base polymorphic marker 1 be a single base polymorphic marker. Table 3 and table 4 said displayed a single base polymorphic marker type can be judges whether skin wrinkles.

Each markers of the present invention determines the frequency of single base polymorphic marker was judged by measuring a number of diagnostic skin type. The significance of the 0. 05 less than, 0. 01 less than, 0. 001 less than, 0. 0001 less than, 0. 00001 less than, 0. 000001 less than, 0. 0000001 less than, 0. 00000001 less than, or 0. 000000001 p-a value of less than one the number such as one characterized 802.11a packets not p - value. Preferably p a-value is 0. 01 can be less than, more preferably p a-value is 0. 001 can be less than disclosed.

Table 3 and table 4 polymorphic marker display as a marker of the present invention, polymorphic marker of an individual gene (minor allele) table 3 and table 4 alleles in the minority base areas when the tube is corrugated (control) is displayed on many skin is skin wrinkles and showed a frequency of high frequency skin test group (case) be dissolved and decides which of the group consisting of, table 3 and table 4 multiple gene alleles (major allele) base areas when displayed on a test group showed many skin is skin wrinkles is corrugated and is dissolved at low frequency marker that can be notified to a user be frequency group of skin are disclosed. For example, when the second base is set forth in table 3 of 7 times of an individual chromosome 24551208 G herein are gene alleles from higher frequency of said regulated since many wrinkles in the skin of an individual in a few G-capturing troops can be judged skin are disclosed. If multiple microarray gene of chromosome 7 times of an individual set forth in table 3 when the registered test group in A 24551208 alleles are skin wrinkles is lower frequency multi-axis can be stored test troops are disclosed.

More preferably, said second base is A or G (rs2711118) in human 7 of table 3 once 24551208 chromosome, including 5 - 100 said 24551208 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 5 76839279 A or G in (rs4273585), including 5 - 100 said 76839279 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 1 208672298 C or T in (rs11119496), including 5 - 100 said 208672298 microarray of continuous DNA sequences polynucleotides; at second base is T or C 52738724 of human chromosome 20 in (rs6127255), including 5 - 100 said 52738724 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 7 27346024 C or T in (rs2158769), including 5 - 100 said 27346024 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 15 47954398 A or C in (rs16963008), including 5 - 100 said 47954398 microarray of continuous DNA sequences polynucleotides; at second base is T or C 101951190 of human chromosome 3 in (rs1048364), including 5 - 100 said 101951190 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 3 101936968 G or A in (rs931206), said 101936968 including 5 - 100 microarray of continuous DNA sequences polynucleotides; at second base is T or 999000 0212999 106740691 of human chromosome 13 in (rs9555341), including 5 - 100 said 106740691 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 10 122472521 C or T in (rs10886747), including 5 - 100 said 122472521 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 4 177330499 G or T in (rs4538426), including 5 - 100 said 177330499 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 4 177322010 C or T in (rs2170576), including 5 - 100 said 177322010 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 12 51222720 G or A in (rs11170170), said 51222720 including 5 - 100 microarray of continuous DNA sequences polynucleotides; at second base is T or C 107123424 of human chromosome 6 in (rs3747790), including 5 - 100 said 107123424 microarray of continuous DNA sequences polynucleotides; at second base is T or G 68751073 of human chromosome 11 in (rs7931342), including 5 - 100 said 68751073 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 1 199554152 C or T in (rs916398), said 199554152 including 5 - 100 microarray of continuous DNA sequences polynucleotides; at second base is of human chromosome 10 in G or A 72820331 (rs10999801), said 72820331 continuous DNA microarray including 5 - 100 of human chromosome 10 times 72805925 9990000289 999 polynucleotides sequences of second base is A or G in (rs1417210), including 5 - 100 said 72805925 microarray of continuous DNA sequences polynucleotides; and complementary polynucleotide sequence consisting of at least one selected from the group consisting of the data, there is provided a skin fold-skin type is determining whether cognitive or corrugated skin wrinkle skin type be a marker for the diagnosis.

In the embodiment of the present invention according to one, many people and the skin wrinkles on human skin wrinkles on the aimed gene using saliva so as to statistically significant SNP 18 of table 3 during inspection of adaptor protein family is one of ABI gene family member 3, involved in oligonucleotides (cell motility) and mobile control (migration regulation) is known. This SNP is phenomena when seen through the viewer appears much wrinkles in assays, high mobility associated with oligonucleotides and pleated egcg appear to substrate. (TRK-a fused gene) TFG does not well known, but which interact with PLSCR 1 is known. Recent studies report that it is involved in TFG NF-a kappa B path (pathway) flow tides. PLSCR 1 (phospholipid scramblase 1) as phospholipid scrambling to number 1, as the number of scrambling can be converted indirectly through the role of TFG. As membrane phospholipids to number scrambling pattern to proteins that are involved (phospholipid translocation), mitochondrial metabolism, lipid metabolism, thrombosis (thrombosis), and apoptosis involved in substrate. As the number is highly TFG interacts with the skin wrinkles scrambling up to automatically control the level of cell apoptosis is through a skin wrinkle can affect the object when supporting each other.

The present invention is demonstrated by the victims of the SNP marker for diagnosing skin type such as confirmed through next. Specifically, the cosmetic skin wrinkle evaluation method (reference KFDA) (visual assessment; VA) functional visual evaluation criteria based on wrinkles VA grade 8 or more of the many controls (control), the tube is corrugated to test group (case) was classification is VA grade 6 hereinafter. The gDNA from blood extracts said classification saliva or from multiple individuals, Axiom experiments (TM) for analyzing gene (also 1) conducting to ASI array plate. As a result, classification of wrinkles (table 3 and table 4) was significantly different skin type with a SNP. The fold-skin type can be classification whether the present invention been solved for the first time by users is SNP are disclosed.

In another aspect, the present invention refers to said detect such markers for the diagnosis of the wrinkles of skin type capable of including a probe or amplifying a number number, composition for the diagnosis of the wrinkles of skin type number under public affairs substrate.

In the present invention terms, "detect such markers for the diagnosis of the wrinkles of skin type probe" areas such as said gene is specifically hybridization reactions by selecting a diagnosis whether skin type wrinkles which means a composition, the method of the specific special number one free gene analysis, the invention is provided to all the known gene can be by detecting method.

In the present invention terms, "wrinkle skin type to amplify the marker for the diagnosis of a number number" such as gene amplification by selecting areas provide the means by which said skin type means a composition for diagnosing whether wrinkles and, preferably said wrinkle skin type marker for the diagnosis of a polynucleotide of a gene amplification method which enables to big.

said polymorphic marker antibody amplification, of suitable buffer under suitable conditions (for example, 4 different nucleoside triphosphate and DNA, RNA polymer such as a number or reverse transcriptase as number) and suitable temperature and mold - DNA synthesis of single stranded oligonucleotides can act starting point indicating said substrate. Said primer suitable length depending on the intended use but, typically 15 to 30 nucleotide sequence disclosed. The mold and the short primer molecules generally require lower temperatures in order to form a stable hybrid. The mold and the primer sequence is completely complementary but are not necessarily, the mold and the complementary hybridization enough to S7 substrate.

In the present invention terms, "primer" have short free 3 'end hydroxyl radicals (free 3' hydroxyl group) having complementary base pair sequence template (template) (base pair) to form nascent strand radiation can be short sequences by big starting point function. Appropriate buffer solution at temperature and polymerization primer (i.e., DNA polymer produced or reverse transcriptase) 4 in the presence of nucleoside triphosphate is different reagents and for DNA synthesis can be disclosure. PCR amplification to predict whether desired product embodiment can be generated through skin type. PCR conditions, the contrast based on the length of the antisense primer into a sense can be publicly known.

gun [lu[lu] Oh this [thu[thu] which it gets torn to quickly reduce the solid support method of the present invention probes or primers, or other widely publicly known method is using can be chemically synthesized. Such nucleic acid sequences in addition the field capable of many publicly known means for automatically. These modified examples but - number of methylation, "cap and", to one or more natural nucleotide analogs substituted, modified between and nucleotide, for example, charged connecting body (example: methyl phosphonate, phosphate gun tree ester, esters capable of inhibiting the formation, such as carbamate) or charged connecting body (example: phosphorothioate, such as phosphor with D thio polycarbonate) changing to 2000.

Another aspect, kits for the diagnosis of the wrinkles of skin type including composition for the diagnosis of the wrinkles of the present invention refers to said skin type number under public affairs substrate.

Said RT-a PCR kit be a kit or DNA chip kit.

SNP marker for diagnosing skin type amplification kit of the present invention identifying or through a polymorphic marker, SNP polymorphic marker identifying the expression level of the mRNA expression levels of diagnosis of skin type can be. Specific as an example, mRNA expression levels of marker for diagnosing skin type in the present invention for measuring kit for performing RT-a PCR kit including the requisite element can be. RT-a PCR kit, each marker for diagnosing skin type of gene specific primer RT-a PCR kit in addition test tubes or other suitable container, reaction buffer (pH various and magnesium concentration), the jade city new the [ley[ley] five tie [tu[tu] which will grow (dNTPs), as the number and reverse it buys the enzyme and enzyme such as Taq - polymer, DNase, RNAse billion number number, be DEPC - (DEPC-a water), sterilization can be like. In addition quantitative regulated gene specific primer pairs can be used. In addition preferably, the kits of the present invention for performing DNA chip kit for diagnosing skin type including be a requisite element. DNA chip kit, generally flat solid support plate, typically a microscope slide is not greater than a nucleic acid species lattice-like arrangement (gridded array) that is attached to a glass surface, a nucleic acid chip surface arranged constant, DNA chip on the chip surface treated nucleic acid complementary nucleic acids contained in multiple hybridization (hybridization) tank prior to massively parallel analysis tool are disclosed.

In another aspect, the present invention refers to said wrinkle skin type including microarray for the diagnosis marker for the diagnosis of a polynucleotide of the wrinkles of skin type number under public affairs substrate.

The DNA or RNA polynucleotide including said microarray may be disclosed. Said probe polynucleotides of the present invention including a polynucleotide microarray under the outside number and consisting of conventional microarray.

To a microarray substrate probe polynucleotides immobilized on a number bath method contrast is well known. Hybridization probe polynucleotides can be polynucleotide in said means, complementary strand nucleic acid capable of binding to sequence-specific oligonucleotide big. The probe of the present invention gene allele specific probes as, during two members such as species derived from nucleic acid fragment with the presence of areas, to modulate the derived from DNA fragments is one hybridization, hybridization is not only derived from other members. In this case hybridization condition allows gene show significant difference in hybridization intensity between alleles, sufficiently stringent hybridization only one of accomplishing gene alleles. The difference between the other form good hybridization can be causing gene alleles. Detecting method of the present invention said probe can be used for diagnosing skin type gene alleles. Said diagnostic method is based on nucleic acid hybridization detection method such as line [pul[pul] unit are included in a, DNA in DNA chip substrate using the same method which was previously form ball number can also be disclosed. Said hybrid microbial rigorous conditions, for example 1M hereinafter salt concentration and thus can be performed usually 25 °C or more. For example, 5x SSPE (750 mm NaCl, 50 mm Na Phosphate, 5 mm EDTA, pH 7. 4) and 2530 °C gene allele specific probes hybridization condition can be suitable.

Skin diagnosis of the present invention immobilized on a substrate of an probe polynucleotides associated with these conventional techniques are used to generate high pressure liquid coolant coil and in addition can be hereinafter for the number. In addition, the detection of nucleic acid hybridization and contrast on microarray hybridization result well known. Said detection for example, nucleic acid sample including the phosphor for example Cy3 and capable of generating a detectable signal of a substance such as Cy5 marker labeling material next, and electroluminescent element from said microarray hybridization on hybridization by a detection can detect the signals.

In another aspect, the present invention refers to obtain DNA from a sample taken from separating from multiple individuals (a); (b) said step (a) amplifying said DNA obtained from single base polymorphic marker or hybridization probe and the areas; and (c) said step (b) the amplified or hybridize confirming base including polymorphic site, the wrinkles of skin type a number of whether ball method number under public affairs substrate.

Terms of the present invention, provide the means by which "individual" fold-skin type whether big subject for diagnosis. Said kit for a hair in, urine, blood, various body fluids, isolated tissue, such as from a DNA sample to obtain isolated cells or with saliva but, the one number are not disclosed.

Said step (a) is any method known to one skilled fulfillment method obtained genome DNA available disclosed.

(B) obtained in step (a) said polymorphic marker DNA amplification or hybridization probe from single base said areas includes any method known to one skilled fulfillment available disclosed. For example, target nucleic acids by PCR by amplifying through localized number can be achieved. Chain reaction (LCR) doped silicon layer and other number (Wu and Wallace, Genomics 4, 560 (1989), such as Landegren, Science 241, 1077 (1988)), transcription amplification (transcription amplification) (such as Kwoh, Proc. Natl. Acad. Sci. USA 86, 1173 (1989)) and self maintaining sequence number is desired (such as Guatelli, Proc. Natl. Acad. Sci. USA 87, 1874 (1990)) based on sequence amplification (NASBA) nucleic acids can be used.

During step (c) of said method comprises determining areas base sequencing analysis, microarray (microarray) by hybridization, gene (allele specific PCR) allele specific PCR, hybridization techniques (dynamic allele-a specifichybridization, DASH) dynamic allele gene, extending PCR analysis, SSCP, TaqMan PCR-a RFLP analysis or techniques, SNPlex platform (Applied Biosystems), mass spectrometry (e.g., of Sequenom MassARRAY system), mini - (mini-a sequencing) sequencing method, system (BioRad) Bio-a Plex, CEQ and SNPstream system (Beckman), Molecular Inversion Probe array technology (for example, Affymetrix GeneChip), and BeadArray Technologies (for example, Illumina GoldenGate and Infinium assays) include, not limited. Said method or other method available to one skilled in the art is provided by the present invention, a sequence consisting, including SNP or other polymorphic marker, one or more an allele of a polymorphic marker gene can be identified. The base areas determining is preferably via SNP chip can be.

Terms in the present invention, a fixing device is "SNP chip" SNP DNA microarray can be confirmation once each base one big.

The TaqMan method (1) DNA fragment amplification primers and TaqMan probe design and number desired to small step; (2) different gene alleles FAM VIC dye labeled with the dyes and probes according (Applied Biosystems); (3) and said DNA as templates, of performing PCR using said primers and probe; (4) of said PCR reaction is completed after, nucleic acid analyzer analysis and verifying the TaqMan analysis plate; and (5) said analysis results from step (1) of poly new the [ley[ley] five mote which will grow determining gene type comprising the following steps.

In said, sequencing analysis can be bio-sequence listing determination using a conventional method, can be performed using automated gene analyzer. In addition, the SNP allele specific PCR gene located on the base 3' primer set comprising said SNP located distally to the device primer PCR method for amplifying an DNA fragments means other. The principle of the method, e.g., when a specific base is substituted in A G, said A 3 'end base including primers and of the relevant size DNA fragments by PCR amplification primer in opposite directions towards the device when performing an electrochemical reaction, the strong base is normally performed when said SNP position A is amplification reactions of band is desired location can be observed, when said base is substituted with G DNA primer complementary mold capable of binding but, 3' end towards complementary binding by amplification reaction is performed as using the fact not number are disclosed. DASH is conventional method can be performed, preferably [phu[phu] phosphorus [su[su] etc. can be performed by method.

On the other hand, PCR analysis is first positioning base nucleotide sequence including DNA fragments extending amplification primer pairs then, all nucleotide added to the deprotection reaction with inactivated by phosphorylation, specific SNP herein extending primer, dNTP mixture, FDD oxy nucleotide, DNA polymerase reaction buffer solution and added to a primer extension reaction by performing combustion chamber. The, base extending primer comprises a SNP located 5 'direction immediately adjacent the base 3' and three of the tail end, and having in addition a nucleic acid encoding the same number base dNTP mixture contains error oxy polynucleotides and polypeptides, polynucleotides may be selected in one SNP base representing said oxy error types. For example, when in G to A replacement, dGTP, dCTP and TTP ddATP when added to the reaction mixture, said base being extended by DNA polymerase in primer comprises a substituted occurs and, after several base is shown in position by primer extension reaction becomes ddATP A base end through the body. If said replacement does not occur, the reaction terminates extending position, extending the length of said primer SNP by comparing representing base for identifying the type is equal to or higher.

The, detecting fluorescent labeling method include extending primer or general bio-sequence listing if used in a determination FDD oxy polynucleotide gene analyzer (e.g., such as ABI yarn Model 3700) can be detected by fluorescence detection using said SNP, - free labeled extension primers and FDD oxy polynucleotide is fixed (matrix assisted laser desorption ionization-a time of flight) MALDI-a TOF estimating method by measuring the molecular weight can be detecting said SNP.

Preferably, the method of the present invention ball number information (d) (minor allele) gene polymorphic site amplified or hybridize when table 3 and table 4 alleles represent prime displayed on a test group showed many skin is skin wrinkles is corrugated and is dissolved at high frequency frequency be skin and decides which of the group consisting of, amplified or hybridize with multiple polymorphic sites (major allele) table 3 and table 4 gene alleles when the base is corrugated and is displayed on the test group showed many skin wrinkles skin is skin further including determining frequency be dissolved at low frequency group may be disclosed.