CN1519330A - Polymorphism of first exon in ADAMTSI gene - Google Patents
Polymorphism of first exon in ADAMTSI gene Download PDFInfo
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- CN1519330A CN1519330A CNA031150241A CN03115024A CN1519330A CN 1519330 A CN1519330 A CN 1519330A CN A031150241 A CNA031150241 A CN A031150241A CN 03115024 A CN03115024 A CN 03115024A CN 1519330 A CN1519330 A CN 1519330A
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Abstract
A single nucleotide polymorphism (SNP) for the extron No.1 of ADAMTS1 gene and a method for analyzing the SNP and activity of said extron No.1 are disclosed. Said SNP is the G->C polymorphism at site No.322 in its sequence shown by SEQ ID No.1 and can cause the change Ala(A)->Pro(P) at the site No.108 of coding protein, so changing protein activity.
Description
Technical field
The present invention relates to the single nucleotide polymorphism of ADAMTS1 gene exon 1.The invention still further relates to the method for the allelic mutation of analyzing the ADAMTS1 gene exon 1.
Background technology
ADAMTS1 is a kind of secretory protein, and the N end has signal peptide, zinc binding site, and the zone of being rich in halfcystine.The ADAMTS protein family all works in the proteolysis process.More have the investigator to find, no matter body result interior, in vitro tests shows that ADAMTS1 has destroyed the vascularization process very effectively, effect is all given prominence to than Thrombospondin-1, endostatin and ADAMTS8.(J.Biol.Chem.274：23349-23357，1999.)。
Before the application, also there is not the relevant report of ADAMTS1 gene exon 1 polymorphism.
Summary of the invention
Purpose of the present invention just provides the polymorphism and the detection method thereof of ADAMTS1 gene exon 1.
In a first aspect of the present invention, a kind of method of single nucleotide polymorphism of the people of detection ADAMTS1 gene exon 1 is provided, it comprises step:
(a) determine the 322nd Nucleotide in sequence shown in the SEQ ID of the people ADAMTS1 gene exon 1 NO:1;
(b) detect whether there is single nucleotide polymorphism in described position.
In a preference, described single nucleotide polymorphism is to have C at the 322nd.
In a second aspect of the present invention, a kind of isolating nucleic acid is provided, it has the sequence shown in the SEQ ID NO:1, and the 322nd is C.
In a third aspect of the present invention, a kind of allele specific nucleic acid primer is provided, its length is 15-50bp, and hybridizes specifically and amplify shown in the SEQ IDNO:1 that contains people ADAMTS1 gene exon 1 the amplified production of the 322nd single nucleotide polymorphism in the sequence.
In a fourth aspect of the present invention, a kind of allele specific oligonucleotide probe is provided, its length is 15-50bp, and hybridizes specifically and detect shown in the SEQ IDNO:1 that contains people ADAMTS1 gene exon 1 the 322nd single nucleotide polymorphism in the sequence.
A kind of test kit comprises:
(1) primer of specific amplification people ADAMTS1 gene exon 1 is right,
(2) detect amplified production and compare the reagent that whether exists variation required, the 322nd single nucleotide polymorphism in the sequence shown in the described SEQ ID NO:1 with normal ADAMTS1 gene exon 1.
Another kind of diagnostic kit comprises: the oligonucleotide probe that the primer that the present invention is above-mentioned and/or the present invention are above-mentioned.
Embodiment
The inventor is by extensive and deep research, have been found that the 108th Ala (A) in the coded protein sequence of ADAMTS1 gene exon 1-Pro (P) polymorphic (be last 322 G-of mRNA〉C polymorphism), finished the present invention on this basis.
According to normal people ADAMTS1 gene sequencing result, found this SNP.Find that by analysis this SNP causes the 108th amino acids by Ala (A)-Pro (P).Because this is the amino acid whose displacement of different properties, therefore, can cause the coded protein-active of ADAMTS1 gene exon 1 to change.
The cDNA sequence of ADAMTS1 gene exon 1 is listed in SEQ ID NO:1, and wherein open reading frame is the 1-372 position, and its coded aminoacid sequence is listed in SEQ ID NO:2.The genome sequence of ADAMTS1 gene exon 1 can obtain from GenBank.
Those skilled in the art will appreciate that a large amount of analytical technologies can be used for detecting whether the site exists single nucleotide polymorphism described in the ADAMTS1 gene exon 1.These technology comprise (but being not limited to): dna sequencing, sequencing by hybridization; Enzymatic mispairing cutting, heteroduple analysis, dot blot, oligonucleotide arrays (DNA chip), Mini-sequencing, Taqman technology, molecular beacon etc., sex change high performance liquid chromatography (DHPLC).
On the other hand, detection method of the present invention is used to assess the susceptibility of the individual ADAMTS1 of suffering from gene exon 1 relative disease.
Being used for specimen of the present invention and being not particularly limited, for detecting SNP, can be DNA or the mRNA that extracting goes out from samples such as blood, tissue.For detect the ADAMTS1 gene exon 1 active for, can anyly contain the sample of ADAMTS1 gene exon 1, as blood etc.
The primer and the probe that are used for the inventive method or detection kit design according to the cDNA sequence (sequence of SEQ ID NO:1) or the genome sequence of ADAMTS1 gene exon 1, and the synthetic technology of available routine is synthesized and got final product.
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 normal condition, people such as Sambrook for example, molecular cloning: laboratory manual (New York:ColdSpring Harbor Laboratory Press, 1989) condition described in, or the condition of advising according to manufacturer.
Embodiment 1
The extracting of gene and order-checking
Extract DNA with conventional phenol chloroform method from people's blood, concentration correction is used for conventional pcr amplification to 20ng/ul.Primer is:
Adopted primer is arranged: 5 '-ctggcgcact gcttctactc-3 ' (SEQ ID NO:3)
Antisense primer: 5 '-tgaagtcgcg tgggatagat a 3 ' (SEQ ID NO:4)
The product of the 445bp that amplifies carries out dna sequencing behind the purifying.
Embodiment 2
The acquisition of SNP
The ADAMTS1 gene is directly checked order.Each sample sequence of measuring is compared, thereby draw the difference of sequence, obtain SNP.
Embodiment 3
Detection kit
Preparation one detects the detection kit of ADAMTS1 gene-correlation disease susceptibility, and wherein, it is right to contain the following primer that amplifies 322 SNP:
Adopted primer is arranged: 5 '-ctggcgcact gcttctactc-3 ' (SEQ ID NO:3)
Antisense primer: 5 '-tgaagtcgcg tgggatagat a-3 ' (SEQ ID NO:4)
Amplified production and normal control are carried out stratographic analysis with sex change high performance liquid chromatograph (DHPLC), can detect 322 G-easily C type SNP.
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?LISTING
＜110〉Research Center of Shanghai Human Genome
＜120〉ADAMTS1 gene exon 1 polymorphism
<130>030115
<160>4
<170>PatentIn?version?3.1
<210>1
<211>445
<212>DNA
<213>Homo?sapiens
<220>
<221>CDS
<222>(1)..(372)
<223>
<400>1
ctg?gcg?cac?tgc?ttc?tac?tcc?ggc?acc?gtg?aat?ggc?gat?ccc?agc?tcg 48
Leu?Ala?His?Cys?Phe?Tyr?Ser?Gly?Thr?Val?Asn?Gly?Asp?Pro?Ser?Ser
1 5 10 15
gct?gcc?gcc?ctc?agc?ctc?tgc?gag?ggc?gtg?cgc?ggc?gcc?ttc?tac?ctg 96
Ala?Ala?Ala?Leu?Ser?Leu?Cys?Glu?Gly?Val?Arg?Gly?Ala?Phe?Tyr?Leu
20 25 30
ctg?ggg?gag?gcg?tat?ttc?atc?cag?ccg?ctg?ccc?gcc?gcc?agc?gag?cgc 144
Leu?Gly?Glu?Ala?Tyr?Phe?Ile?Gln?Pro?Leu?Pro?Ala?Ala?Ser?Glu?Arg
35 40 45
ctc?gcc?acc?gcc?gcc?cca?ggg?gag?aag?ccg?ccg?gca?cca?cta?cag?ttc 192
Leu?Ala?Thr?Ala?Ala?Pro?Gly?Glu?Lys?Pro?Pro?Ala?Pro?Leu?Gln?Phe
50 55 60
cac?ctc?ctg?cgg?cgg?aat?cgg?cag?ggc?gac?gtc?ggc?ggc?acg?tgc?ggg 240
His?Leu?Leu?Arg?Arg?Asn?Arg?Gln?Gly?Asp?Val?Gly?Gly?Thr?Cys?Gly
65 70 75 80
gtc?gtg?gac?gac?gag?ccc?cgg?ccg?act?ggg?aaa?gcg?gag?acc?gaa?gac 288
Val?Val?Asp?Asp?Glu?Pro?Arg?Pro?Thr?Gly?Lys?Ala?Glu?Thr?Glu?Asp
85 90 95
gag?gac?gaa?ggg?act?gag?ggc?gag?gac?gaa?ggg?gct?cag?tgg?tcg?ccg 336
Glu?Asp?Glu?Gly?Thr?Glu?Gly?Glu?Asp?Glu?Gly?Ala?Gln?Trp?Ser?Pro
100 105 110
cag?gac?ccg?gca?ctg?caa?ggc?gta?gga?cag?ccc?aca?ggtagaagaa 382
Gln?Asp?Pro?Ala?Leu?Gln?Gly?Val?Gly?Gln?Pro?Thr
115 120
ttgctctcgt?ttgtctgtcc?attcatcctc?ctctcctcac?tttatctatc?ccacgcgact 442
tca 445
<210>2
<211>124
<212>PRT
<213>Homo?sapiens
<400>2
Leu?Ala?His?Cys?Phe?Tyr?Ser?Gly?Thr?Val?Asn?Gly?Asp?Pro?Ser?Ser
1 5 10 15
Ala?Ala?Ala?Leu?Ser?Leu?Cys?Glu?Gly?Val?Arg?Gly?Ala?Phe?Tyr?Leu
20 25 30
Leu?Gly?Glu?Ala?Tyr?Phe?Ile?Gln?Pro?Leu?Pro?Ala?Ala?Ser?Glu?Arg
35 40 45
Leu?Ala?Thr?Ala?Ala?Pro?Gly?Glu?Lys?Pro?Pro?Ala?Pro?Leu?Gln?Phe
50 55 60
His?Leu?Leu?Arg?Arg?Asn?Arg?Gln?Gly?Asp?Val?Gly?Gly?Thr?Cys?Gly
65 70 75 80
Val?Val?Asp?Asp?Glu?Pro?Arg?Pro?Thr?Gly?Lys?Ala?Glu?Thr?Glu?Asp
85 90 95
Glu?Asp?Glu?Gly?Thr?Glu?Gly?Glu?Asp?Glu?Gly?Ala?Gln?Trp?Ser?Pro
100 105 110
Gln?Asp?Pro?Ala?Leu?Gln?Gly?Val?Gly?Gln?Pro?Thr
115 120
<210>3
<211>20
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
＜223〉primer
<400>3
ctggcgcact?gcttctactc 20
<210>4
<211>21
<212>DNA
＜213〉artificial sequence
<220>
<221>misc_feature
＜223〉primer
<400>4
tgaagtcgcg?tgggatagat?a 21
Claims (7)
1. method that detects the single nucleotide polymorphism of people ADAMTS1 gene exon 1 is characterized in that it comprises step:
(a) determine the 322nd Nucleotide in sequence shown in the SEQ ID of the people ADAMTS1 gene exon 1 NO:1;
(b) detect whether there is single nucleotide polymorphism in described position.
2. the method for claim 1 is characterized in that, described single nucleotide polymorphism is to have C at the 322nd.
3. an isolating nucleic acid is characterized in that, it has the sequence shown in the SEQ ID NO:1, and the 322nd is C.
4. allele specific nucleic acid primer, it is characterized in that, its length is 15-50bp, and hybridizes specifically and amplify shown in the SEQ ID NO:1 that contains people ADAMTS1 gene exon 1 the amplified production of the 322nd single nucleotide polymorphism in the sequence.
5. an allele specific oligonucleotide probe is characterized in that, its length is 15-50bp, and hybridizes specifically and detect shown in the SEQ ID NO:1 that contains people ADAMTS1 gene exon 1 the 322nd single nucleotide polymorphism in the sequence.
6. diagnostic kit is characterized in that it comprises:
(1) primer of specific amplification people ADAMTS1 gene exon 1 is right,
(2) detect amplified production and compare the reagent that whether exists variation required, the 322nd single nucleotide polymorphism in the sequence shown in the described SEQ ID NO:1 with normal ADAMTS1 gene exon 1.
7. a diagnostic kit is characterized in that it comprises: described primer of claim 4 and/or the described oligonucleotide probe of claim 5.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA031150241A CN1519330A (en) | 2003-01-22 | 2003-01-22 | Polymorphism of first exon in ADAMTSI gene |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA031150241A CN1519330A (en) | 2003-01-22 | 2003-01-22 | Polymorphism of first exon in ADAMTSI gene |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN1519330A true CN1519330A (en) | 2004-08-11 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNA031150241A Pending CN1519330A (en) | 2003-01-22 | 2003-01-22 | Polymorphism of first exon in ADAMTSI gene |
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| CN (1) | CN1519330A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2014011997A (en) * | 2013-07-16 | 2014-01-23 | Univ Of Tokyo | Osteoporosis susceptibility gene, and method for determination of risk of acquiring osteoporosis |
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2003
- 2003-01-22 CN CNA031150241A patent/CN1519330A/en active Pending
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2014011997A (en) * | 2013-07-16 | 2014-01-23 | Univ Of Tokyo | Osteoporosis susceptibility gene, and method for determination of risk of acquiring osteoporosis |
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