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PNAClamp 技術(shù)是指PNA能夠?qū)Ｒ恍缘呐c野生型DNA結(jié)合，從而使野生型DNA不能作為模板進(jìn)行PCR擴(kuò)增；而突變型DNA，由于堿基的缺失或突變，PNA不能與其結(jié)合，從而使突變型DNA能夠進(jìn)行PCR擴(kuò)增。
一般來說，一個(gè)樣本中，大部分是野生型DNA，突變型DNA只占極少的一部分，因此通過一般的PCR很難檢測到突變型DNA的存在。如果在樣本中加入與野生型DNA互補(bǔ)的PNA，則野生型DNA與PNA牢固結(jié)合，使得野生型DNA不能進(jìn)行PCR擴(kuò)增，由于沒有野生型DNA的干擾，突變性DNA能夠很容易被擴(kuò)增并檢測。
PNAClamp Mutation detection 的原理：
1. 與 DNA/DNA 的結(jié)合相比，在不存在錯(cuò)配的情況下，PNA/DNA 的結(jié)合穩(wěn)定性更強(qiáng)（每增加 1 base 序列，退火溫度增加 ～1℃）。
2. 與 DNA/DNA 的結(jié)合相比，在存在錯(cuò)配的情況下，PNA/DNA 的結(jié)合穩(wěn)定性更差。
3. PNA 不能作為引物被 DNA polymerase 識別并擴(kuò)增。
PNAClamp Mutation detection Kits 的優(yōu)勢：
1. 檢測時(shí)間短（約2小時(shí)）。
2. 操作簡單。
3. 需要樣品少（低至1～25ng DNA樣品）。
4. 靈敏度高。
5. 穩(wěn)定性強(qiáng)。
6. 保質(zhì)期長。
7. 可以在多種型號的熒光定量PCR儀中使用（如：BI-7500, ABI-7900HT, Biorad CFX-96/384, Roche Lightcycler, Qiagen RGQ etc）。
8. 數(shù)據(jù)分析簡單明了。
PNAClamp Mutation detection 示意圖：
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The epidermal growth factor receptor (EGFR) is a family member of Receptor tyrosine kinases, expressed on the surface of epidermal cells. Overexpression or overactivation of EGFR is linked to a number of cancers, including lung cancer, anal cancers and glioblastoma multiform.
The PNAClamp EGFR Mutation Detection Kit detects most prevalent mutations described to date in the EGFR gene, including T790M, the presence of which correlates with resistance to tyrosine kinase inhibitors. Detecting somatic mutations in EGFR gene may provide a useful strategy to predict the response to the tyrosine kinase inhibitors in efforts to increase the survival rate of lung cancer patients receiving targeted therapy.
Exon
Amino acid change
Base change
18
Gly719Ala
2156 G>C
Gly719Ser
2155 G>A
Gly719Cys
2155 G>T
19
Glu746_Ala750del
2235_2249 del 15
Glu746_Thr751delinslle
2235_2252 AAT (complex)
Glu746_Ser752del
2236_2253 del 18
Glu746_Thr751delinsAla
2237_2251 del 15
E746_S752>A
2237_2254 del 18
Glu746_Ser752delinsVal
2237_2255 >T (complex)
Glu746_Ala750del
2236_2250 del 15
Glu746_Ser752delinsAsp
2238_2255 del 18
L747_A750>P
2238_2248 >GC (complex)
Leu747_Thr751delinsGln
2238_2252 >GCA (complex)
Leu747_Glu749del
2239_2247 del 9
Leu747_Thr751del
2239_2253 del 15
Leu747_Ser752del
2239_2256 del 18
Leu747_Glu749del:Ala750Pro
2239_2248 TTAAGAGAAG>C
Leu747_Pro753delinsGln
2239_2258 >CA (complex)
Leu747_Thr751delinsSer
2240_2251 del 12
Leu747_Pro753delinsSer
2240_2257 del 18
Leu747_Thr751del
2240_2254 del 15
Leu747_Thr751deinsPro
2239_2251 >C (complex)
20
Thr790Met
2369 C>T
Ser768lle
2303 G>T
Ala767_Val769dupAlaSerVal
2307_2308 ins9
His773dupHis
2319_2320 insCAC
Asp770_A771insGly
2310_2311 insGGT
21
leu858Arg
2573 T>G
leu861Gln
2582 T>A
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KRAS mutation is found in several cancers including colorectal, lung, thyroid, and pancreatic cancers and cholangiocarcinoma. KRAS mutations are often located within codons 12 and 13 of exon 2, which may lead to abnormal growth signaling by the p21-ras protein. These alterations in cell growth and division may trigger cancer development as signaling is excessive.
A KRAS mutation often serves as a useful prognostic marker of drug response. For example, a KRAS mutation is considered to be a strong prognostic marker of response to tyrosine kinase inhibitors such as gefitinib (Iressa) or erlotinib (Tarceva). Recently, KRAS mutations have been detected in many colorectal cancer patients and may be associated with responses to cetuximab (Erbitux) or panitumumab (Vectibix), which are used in colon cancer therapy.
Codon
Mutation
Base change
Codon12
Gly12Asp
35G>A
Gly12Ala
35G>C
Gly12Val
35G>T
Gly12Ser
34G>A
Gly12Arg
34G>C
Gly12Cys
34G>T
Codon13
Gly13Ser
34G>T
Gly13Arg
37G>C
Gly13Cys
37G>T
Gly13Asp
38G>A
Gly13Ala
38G>C
Gly13Val
38G>T
Codon59
Ala59Ser
175G>T
Ala59Thr
175GA
Ala59Glu
176C>A
Ala59Gly
176C>G
Ala59del
176_178 del CAG
Codon61
Gly60Asp
179G>A
Gly60Ala
179G>C
Gly60Val
179G>T
Gly60Gly
180T>A
Gly60Gly
180T>C
Gln61Glu
181C>G
Gln61Lys
181C>A
Gln61Leu
182A>T
Gln61Arg
182A>G
Gln61Pro
182A>C
Gln61His
183A>T
Gln61His
183A>C
Codon 117
Lys117Glu
349A>G
Lys117Arg
350A>G
Lys117Asn
351A>C
Lys117Asn
351A>T
Codon 146
Ala146Pro
436G>C
Ala146Thr
436G>A
Ala146Gly
437C>G
Ala146Val
437C>T
Ala146Ala
438A>G
Ala146Ala
438A>C
Ala146Ala
438A>T
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PI3K (Phosphoinositide 3-kinases or PI 3-Kinases) are family of lipid kinases capable of phosphorylating the 3' position hydroxyl group of the inositol ring of phosphatidylinositol. They are involved in coordinating a diverse range of cell functions including cell growth, proliferation, differentiation, motility, survival and intracellular trafficking.
Activating mutations in PI3K catalytic domain of the p110alpha subunit (PIK3CA) have recently been discovered in certain types of cancer cells. PIK3CA mutations are found at 25~40 % frequency in various types of tumors including colorectal cancer, gastric cancer, lung cancer, brain cancer, endometrial cancer, ovarian cancer, breast cancer. About 80% of the point mutations presides in exon 9 (a presumed helical domain) and exon 20 (a presumed kinase domain), while other types of mutations are also seen in different locations.
Tube No
Reagent
Amino Acid Change
Base change
Exon
Translation region
1
E542
Glu542Lys
1624 G>A
Exon 9
Helical
Glu542Gly
1624 A>G
Glu542Val
1624 A>T
2
E545
Glu545Lys
1633 G>A
Glu545Gly
1634 A>G
Glu545Asp
1635 G>T
3
H1047
His1047Tyr
3139 C>T
Exon 20
Kinase
His1047Leu
3140 A>T
His1047Arg
3140 A>G
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產(chǎn)品信息：
Catalog No.
Product Name
Description
Size
Price
PNAC-1002
PNAClamp KRAS Kit (v2)
G12, G13
30 tests
PNAC-1003
PNAClamp KRAS Kit (v3)
G12, G13, Q61
25 tests
PNAC-1004
PNAClamp KRAS Kit (plus)
A59, K117, A146
25 tests
PNAC-1006
PNAClamp KRAS Kit (v4)
G12, G13, A59, Q61, K117, A146
25 tests
PNAC-1101
PNAClamp NRAS Kit (v4)
G12, G13, A59, Q61, K117, A146
25 tests
PNAC-2001
PNAClamp BRAF Kit
BRAF V600 mutation
50 tests
PNAC-3002
PNAClamp EGFR Kit
G619, E19 del, T790, S768, E20 in, L858, L861
25 tests
PNAC-4002
PNAClamp PIK3CA Kit
E542, E545, H1047
25 tests
PNAC-5001
PNAClamp IDH1 Kit
R132 mutation
25 tests
PNA Clamp Kit
