KS J 0009-2007 Biotechnology-Standard guide for detection of nucleic acid sequence by the polymerase chain reaction techniquesequence by the polymerase chain reaction technique《生命工.pdf

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1、 KS J 0009 KSKSKSKS SKSKSKS KSKSKS SKSKS KSKS SKS KS (PCR) KS J 0009 : 2007 (2012 ) 2007 10 24 http:/www.kats.go.krKS J 0009：2007 ： ( ) ( ) KT&G ( ) : (http:/www.standard.go.kr) ： ：2002 11 30 ：2007 10 24 ：2012 12 27 ： 20120774 ： ( 02-509-7270) (http:/www.kats.go.kr). 10 5 , . KS J 0009：2007 ii 1 1 2

2、 1 3 .2 4 .5 5 .5 6 (target materials).6 7 PCR 6 8 / 7 9 8 9.1 DNA .8 9.2 RNA PCR(RTPCR) .8 9.3 (dNTPs) .8 9.4 PCR .9 9.5 9 9.6 DNA9 9.7 RNA PCR RNA 9 9.8 DNA 10 9.9 .10 10 PCR .10 10.1 .10 10.2 .10 10.3 /PCR 11 11 PCR .11 11.1 .11 11.2 .12 12 12 13 13 14 13 15 13 A() 15 i KS J 0009：2007 ii (PCR) (D

3、NA) (RNA) . PCR , , DNA . . PCR . , , , , PCR , . . PCR , . KS J 0009 : 2007 (2012 ) (PCR) Biotechnology Standard guide for detection of nucleic acid sequence by the polymerase chain reaction technique 1 1.1 PCR , , , . PCR . . 1.2 PCR DNA RNA . RNA RNA DNA(cDNA) PCR . PCR(RTPCR) . 1.3 / . , , . 1.4

4、 PCR . 1.5 . PCR . 1.6 PCR DNA . 1.7 PCR RTPCR ( PCR, PCR in situ PCR) . 1.8 . 2(Biosafty level 2) . 8 . 2 . KS J 0009：2007 . ( ) . 2.1 NCCLS I17P, Protection of Laboratory Workers from Instrument Biohazards, Proposed Guideline(1991) C24A, Internal Quality Control Testing：Principles and Definitions,

5、 Approved Guideline(1991) M29T2, Protection of Laboratory Workers from Infectious Disease Transmitted by Blood, Body Fluides, and TissueSecond Edition, Tentative Guideline(1991) GP5A, Clinical Laboratory Waste Management：Approved Guideline(1993) NM3A, Molecular Diagonostic Methods for Infectious Dis

6、eases, Approved Guideline(1995) 3 . 3.1 (annealing) DNA RNA (primer) DNA DNA RNA 3.2 (carryover contamination) (PCR ) PCR . . 3.3 (chaotrope) 3.4 (denaturation) DNA DNA . , pH . 3.5 DNA (DNA polymerase) DNA PCR . PCR (cycle) , DNA . 3.6 DNA (DNA probe) 20 35 , DNA . DNA , . DNA PCR . DNA PCR . 2 KS

7、J 0009：2007 3.7 () (extension) DNA DNA DNA . DNA . 3.8 PCR (hot-start PCR) ( ) PCR , . PCR , ( Mg ) . 60 ( ) ( PCR DNA ) . 3.9 (hybridization) pH DNA RNA DNA DNA ( DNA ) 3.10 PCR (nested PCR) PCR PCR. PCR 2 PCR 2 . 1 PCR 2 3.11 (polymerase chain reaction) PCR (1 3). 2 DNA DNA . 3 ( DNA , DNA DNA ) D

8、NA . . 3.12 PCR (PCR buffer) PCR DNA , pH 3.13 PCR (PCR product) PCR DNA , DNA. (amplicon) . 3.14 (primer) DNA 20 30 bp 3 KS J 0009：2007 . DNA . . RT PCR 6 RNA . 3.15 (primerdimer) , 2 3 . . PCR , DNA DNA . 3.16 (reaction tube) 100 4 . PCR PCR . PCR . 2,9,103.17 PCR (reverse transcription PCR, RTPCR

9、) RNA . 3,11 RNA cDNA RNA DNA . cDNA ： RNA DNA cDNA DNA 2 DNA . cDNA () DNA , DNA DNA (3.2.11 ). (DNA) RNA (antisense primer) . 3.18 / (reverse transcriptase/polymerase enzyme) RNA cDNA PCR cDNA . rTth Mn RNA , RNA cDNA DNA . M MVL AMV RNA cDNA , PCR cDNA DNA . 3.19 (target sequence) DNA RNA(RTPCR),

10、 PCR (sources) . 3.20 (thermal cycler) RTPCR RNA PCR DNA . , (well) . 4 KS J 0009：2007 3.21 DNA (thermostable DNA polymerase) PCR DNA DNA . . Taq PCR, . 4 4.1 PCR RTPCR . 4.2 PCR . 5 5.1 PCR . DNA() 2 , , Mg DNA . DNA . 5.1.1 2 DNA . 5.1.2 DNA . 5.1.3 DNA 2 DNA . 5.1.4 (5.1.1 5.1.3) . ( 30). , . 5.2

11、 RTPCR RNA cDNA . RNA cDNA , , 2 . 2 cDNA PCR (). cDNA 2 , , Mg DNA . RNA . 5.2.1 RNA cDNA . 5.2.2 cDNA：RNA . 5.2.3 DNA . 5 KS J 0009：2007 5.2.4 DNA 2 . 5.2.5 5.2.2 5.2.4 . ( 30). 1 DNA 2 . , . 5.3 PCR RT PCR PCR . (blot) (solid phase detection formats), (in situ hybridization), (microtiter wells),

12、(bead), (solution hybridization formats), (exclusion) , (hydroxyapatite) , (affinity) (capturechromatography), (homogeneous solution hybridization with labeled probes), HPLC . , PCR . . 4 7 NCCLS MM3A . 6 (target materials) 6.1 , . DNA , , , PCR . , . . . . (5 ) 4 . 2080 . . DNA TE(TrisEDTA). 6.2 PC

13、R , . , , , , . . / , / . 7 PCR 7.1 DNA DNA 2 . . 6 KS J 0009：2007 7.1.1 (pre-amplification area) . PCR PCR . 7.1.1.1 PCR . 7.1.1.2 . 7.1.2 (post-amplification area) , . 7.1.3 . , . . PCR . . . . . . 8 / 8.1 PCR . 4 99 . 35 99 30 5 . 1 . 8.2 8.3 100 4 PCR . . PCR . 8.4 (light mineral oil) . . . (wel

14、l) . 8.5 DNA , , , , , , , 7 KS J 0009：2007 , HPLC PCR , . 8.6 , , . NCCLS GP 5 . PCR , . PCR 7., , 10.2.2 . 9 9.1 DNA (thermal cycling) PCR . . . PCR . A . PCR . , . . . (lot) , . . 9.2 RNA PCR(RTPCR) DNA . M MLV AMV DNA . . PCR . , . . (lot) . 9.3 (dNTPs) DNA PCR dATP(), dCTP( ), dGTP() TTP() . dUTP PCR . . PCR DNA DNA . . dNTPs . dNTPs , . dNTP 8 KS J 0009：2007 200 M. 9.4 . 9.4 PCR PCR PCR . . . Mg ( rTth Mn ) . pH