技術原理:
數字PCR( 也可稱單分子PCR) 一般包括兩部分內容,即PCR 擴增和熒光信號分析。在PCR 擴增階段,與傳統技術不同,數字PCR
一般需要將樣品稀釋到單分子水平,并平均分配到幾十至幾萬個單元中進行反應。不同于qPCR 對每個循環進行實時熒光測定的方法,數字PCR
技術是在擴增結束后對每個反應單元的熒光信號進行采集。最后通過直接計數或泊松分布公式計算得到樣品的原始濃度或含量。數字PCR
的定量方法不依賴于擴增曲線的循環閾值,因此不受擴增效率的影響,也不必采用看家基因和標準曲線,具有很好的準確度和重現性,可以實現絕對定量分析。在未來十年內,醫學保健的目標是提供優質高效的個性化醫療。PCR
方法會在實現這個目標的過程中發揮重要作用。相比于傳統的PCR,數字PCR 技術有著無可比擬的優勢和廣泛的應用前景。

數字PCR檢測原理
應用領域:
·突變/稀有變異檢測(Mutation/Rare
variant detection)
·拷貝數變異(Copy-number
variation)
·進入外周循環(包括血液和糞便)的腫瘤組織核酸分析
·無創產前篩查
·轉化移植檢測
·藥理學(Pharmacogenetics)
·基因表達分析(Gene expression
analysis)-特別針對單細胞或少量細胞或者血清血漿樣品
·線粒體拷貝數分析與線粒體突變分析
·數字PCR可以驗證二代測序(NGS)
應用案例:
·Huggett
JF, Foy CA, Benes V et al.(2013) The digital MIQE guidelines: minimum
information for publication of quantitative digital PCR experiments. Clinical
Chem 6:892–902.
·Absolute
quantification by droplet digital PCRversus analog real-time PCR Christopher M
Hindson et al. nature methods | VOL.10 NO.10 | OCTOBER 2013.
·Detection
of Cancer DNA in Plasma of Early Stage Breast Cancer Patients Julia A. Beaver*1,
Danijela Jelovac February 19, 2014 American Association for Cancer Research.
·Evaluation
of a Droplet Digital Polymerase Chain Reaction Format for DNA Copy Number
Quantification Leonardo B. Pinheiro, Victoria A. Coleman,November 28, 2011
American Chemical.
·Qin
J, Jones R C, Ramakrishnan R. Studying copy number variations using a
nanofluidic platform. Nucleic Acids Res, 2008,36(18): e116.
·Lo
Y M D, Fiona M F, Chan K C A, et al. Digital PCR for the molecular detection of
fetal chromosomal aneuploidy. Proc Natl Acad Sci USA, 2007, 104(32):
13116-13121.
·Warren
L, David B, Irving L W, et al. Transcription factor profiling in individual
hematopoietic progenitors by digital RT-PCR. Proc Natl Acad Sci USA, 2006,
103(47): 17807-17812.
·Ottesen
E A, Jong W H, Stephen R Q, et al. Microfluidic digital PCR enables multigene
analysis of individual environmental bacteria. Science, 2006, 314(5804):
1464-1467.
·Tadmor
A D, Elizabeth A O, Jared R L, et al. Probing individual environmental bacteria
for viruses by using microfluidic digital PCR. Science, 2011, 333(6038): 58-62.
·Tian
Wang, Hongying Sha. , et al. Polar Body Genome Transfer for Preventing the
Transmission of Inherited Mitochondrial Diseases。Cell 157, 1591–1604, June 19,
2014.
![]() |
![]() |
|
![]() |
![]() |
![]() |
![]() |
![]() |
![]() |
![]() |
![]() |
![]() |