LAMP技术介绍.pdf

1、LAMP0 F ),1y$Y_ (loop-mediated isothermal amplification,LAMP_ 2000+a.D0 Notomi 1y 1 ,0/ ,F1y$Y(2hG(w!,_ B_k?_Y+XT)(!AA,E( 9K.5#kW, DNA 6 8G X QT1E(5 84)*)( K.5 AE( X1y$Y4 65 )NY5 8: F |K.5 =P5 PCR M0?FJE=, WE0;9 K. Q,5j Y+X +O)(5,E( Bst DNA 6 8GK.5 8,#kW XMGT1E(5 84)= +O)( K.5E( X1y$YE K. 86 F2c , F

2、2 GF TZGL$-$-(F 5-F1c -F2-3;BIP 5 B1c(B1 jG ) B2 GT!G-$-(F 5-B1c-B2-3FGE(j6 F3c B3c , F3 B3 G *6LAMP ,E0; 6j9ZLf!) Q 8Lf!)Lf! hK)Lf! 1 X) Q 8Lf!E( FIP j Tm l #G-WFGE( F3 p FIP , F2 Gx F3 5 8 Q DNA , F2c :E, 8, DNA K. . 25 1 3W-pFE( F3 5 8 Q DNA , F3c : . 25 2 E, 8 Q DNA ,K. . 25 3 FJE5 Gn*+a FIP E,

3、8,K. . 25 4 Gn*,K. 51, F1c F1 +O8A.*G)0Z)(5 K. 51,)(502l F3,FE( B3 D-p BIPE(F5 8 B3c :E, 8BK.,K. K. . 25 4 5 5*,K.T16V9 B1c-B2-B1 F1-F2c-F1c 58f +O.*G)0T1)(, J? K.5 . 25 6 B5_ LAMP #* ),C5_ 5) , QX)Lf! J?, K. DNA 5FJE8AE,FJE2l56 4 , . 25 13 14 +O0Z5 6 0g, DNA K. 0Z5 9 , DNA K. . 25 15 6)Z F$ .25 9-1

4、1 0 ,(_03+GW?=0,+a AG=,MG,9)5 J)8: 5, DNA (w!$ 8( . 25 16 17 RT-LAMP F6E LAMP (RT-LAMP)_ X LAMP*.p:*0C, )(2RNA6,FO1y$Y,_ 1, 2 *6 LA MP-( | RT-LAMP, 365nm PEI-DNA= 8(“#*)6-l ?,7A9y6JT3P5 8C)M6JT3P5 8F 0!jJT3P,9y X3F +O DNA ,1 n-;JT3P *,5/; 89y 89 LAMP _ 8 LAMP _ B_J X=, LF6E G4 8) RNA Q,FO# RT-LAMPF-

5、F) X;M7 M W Q , +X m1 , LAMP_ X1y$YvM0?x) Q DNAF | 95 5min P,N W4*6 Kentaro Nagamine1y9)+XM W4*6 DNA QF | LAMP , |WF |3R,.D0 )A_+XV9 HBV DNA(w!,CX2F_d HBV-DNALcW+6/p$5g =F | Q,N W4*60g NYF | LAMPFJE, DNA Q DNA2* 4 DNA1yA=F |N W4*6, |W=M0?C W4*6 LAMP_ Xd+XF |$C $FO,#8 ) ( E( , +X jQ LAMP ,FO Kentaro

6、Nagamine1y 10 X 4E(,*.p: 2M A ,)(5,)E(WWQ LAMP ,FOT)E( F B_FJE9)5r _K.5 DNA, 8vp5 8,)5E(pM A,5=0g)E( F Loop F B(Loop B)5 8, j6 F1 F2L$ B1 B2L$65 8E DNA 86nQ 6+X)E(, LAMP 5# XZE0; X0Z?F |K.5#k6|: /$03 9y r (FISH)_ X8f)#+ ,1p X47,W? +O*6#kW1yF“# +X+ ,#5 8 r_, PCR9x Q#,x) (2Wv |# 47,+X 47 DNA6 8Gf , 1y

7、 Maruyama1y 12 6 LAMP r5 8+X#jV stxA 2* , E coliO157 H7= PCR,5jD-()E,$YF | A)47,.d 9Y+X9yF |PEI“#5 8,9y-#“8#JT3P9y-#“8#1y? *6G6 F 1-$?,#, LAMP_-O!)FOA7) jFO#+X,W% _ ? _ LAMP _,h4.D FZ?J/n“0 *CL+XAA LAMP E(,E8mM MgSO4;0.1% Tween20 0.8M Betaine+L: . ;1.4 mM dNTP;1.6uM FIP 1.6uM BIP 0.2uM F3 0.2uM B3 0

8、.8uM loop-F 0.8uM loop-B 0G, Bst DNA6 8G DNA Q 3+,G+XG E(+XG F |H FB3(_GE(,+XGv AFGE(,# XGE(#, 1/41/10 L$!FE(,4?#?0+X HPLC 4 F,E( LAMP 0; _6 3+5 6065 $Y 3060minf 80PLQ4! F |(# . . #q 0; . 9 ( #LAMP(# 5*l723+e# . 5 A, F5Lf? .Ba 9yG# ( .+*) S “4 ,#zT_ V ( . 5 B) +X6yyA# 400nmy hfF |# Teramecs (0 ,CL+X

9、LAMP(#,#z LAMP 3+005G- X= F | p LAMP )FOQx,Z X 3060min * 1091010=MGCM , DNA GQE500ug/ml(9 ) ( 2WQ 4 ZE(M A 6 Z j LAMP ,Q(2W 6 Z)0,G XCLf!MG,A65, I+a 4Z)0,GMG,A !a1 XMM DNA ; LAMP ,(2G-= a7- FJE) -dg ,# +“0+“ +F |0!#57/$03#B#* FF$W51$6jLcW #+O+,+b #qO+“3 X 3Z? 7A PLQ (FJE#z)LE3)8OW“h?5738 )GJ49.D05P5

10、 PCR -(“ LAMP_=FOO6 636 X B1 SYBR Green F |#(4)-O9 , #Endo1y 34+XLAMP_#$?2*39 (PB) PB_EC 5(+O9+(PCM),C-(-O9 PCMQ 15#b -A“ELP5PCR-O9#* PB(2,* (gp43 gp271y )M0? 56h6p#F447#IM0? 12h: Endo1y*0, LAMP# gp43* 5 DNA XM0 3 hn/j LAMP_ X-O9#M,8 76 W H X7 6.D07 6W,TG? Hirayama1y 35F+X LAMP#) Y8(2GF |#H(7 6W 8x4

11、En7,7674*6 DNAf w6 X 2ZB1 | 2!Q LAMP LW(2, LL9, LAMP +X#LW7 6,(2G S4_ Y8:,0!G=G+XLL9, LAMP ,(2G_ 1.715 ODNAG 2Z LA MP G-jLcW 5 DNA X=CE 1 h(,#I Mh Y+X+O,8“#F |#z6 ! LAMP X7 6WHM9W,+X +7 6+O(0;M,LD2 j 9 SNP #h9GJ1W single nucleotide polymorphisms, SNPsj19FP7A X=r+,* 66Z+COW69(* 4,.D0 U-p88G?,+X FJE-$

12、# K.BJ1W6*8(w#F |#v X4q*#Q1yLNF+X LAMP _ )1, SNP #* *6_6 SNP #zn6-;=#z f LAMP +X SNP#9Q W O 81983OE* =(LFF+| 4 - vJ F/=, 42E* =( “F,/*)E* ( -(O ,WAM0?E* NF |# 7AE* O (GMOs),#08_FJE#E,F$ _TWJ E* O G- 9, CaMV 35S _ LA MP_ &FO1) CJ.A?“r1y(&“F!+XO 6 Shiro1y 36M A CaMV 35S _* *0 LAMP#FJE(V/GKq+O,#zTWBv_

13、VF |E* i8http:/loopamp.eiken.co.jp/e/index.htmlShiro Fukuta,Yuko Mizukami,Akira Ishida,et al., Real-time loop-mediated isothermal amplification for the CaMV-35S promoter as a screening method for genetically modified organisms. Eur Food Res Technol,2004,218:496500. 63 )1. Notomi, T., et al., Loop-me

14、diated isothermal amplification of DNA. Nucleic Acids Res, 2000. 28(12): p. E63. 2. Soliman, H. and M. El-Matbouli, Reverse transcription loop-mediated isothermal amplification (RT-LAMP) for rapid detection of viral hemorrhagic septicaemia virus (VHS).Vet Microbiol, 2006. 114(3-4): p. 205-13. 3. Gun

15、imaladevi, I., et al., A loop mediated isothermal amplification (LAMP) method for detection of infectious hematopoietic necrosis virus (IHNV) in rainbow trout (Oncorhynchus mykiss). Arch Virol, 2005. 150(5): p. 899-909. 4. Mori, Y., et al., Detection of loop-mediated isothermal amplification reactio

16、n by turbidity derived from magnesium pyrophosphate formation. Biochem Biophys Res Commun, 2001. 289(1): p. 150-4. 5. Mori, Y., et al., Real-time turbidimetry of LAMP reaction for quantifying template DNA. J Biochem Biophys Methods, 2004. 59(2): p. 145-57. 6. Yoshida, A., et al., Loop-mediated isoth

17、ermal amplification method for rapid detection of the periodontopathic bacteria Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola. J Clin Microbiol, 2005. 43(5): p. 2418-24. 7. Mori, Y., T. Hirano, and T. Notomi, Sequence specific visual detection of LAMP reactions by addition

18、of cationic polymers. BMC Biotechnol, 2006. 6: p. 3. 8. Blomstrom, A.L., et al., A one-step reverse transcriptase loop-mediated isothermal amplification assay for simple and rapid detection of swine vesicular disease virus. J Virol Methods, 2008. 147(1): p. 188-93. 9. Nagamine, K., et al., Loop-medi

19、ated isothermal amplification reaction using a nondenatured template. Clin Chem, 2001. 47(9): p. 1742-3. 10. Nagamine, K., T. Hase, and T. Notomi, Accelerated reaction by loop-mediated isothermal amplification using loop primers. Mol Cell Probes, 2002. 16(3): p. 223-9. 11. Nagamine, K., Y. Kuzuhara,

20、 and T. Notomi, Isolation of single-stranded DNA from loop-mediated isothermal amplification products. Biochem Biophys Res Commun, 2002. 290(4): p. 1195-8. 12. Maruyama, F., et al., Detection of bacteria carrying the stx2 gene by in situ loop-mediated isothermal amplification. Appl Environ Microbiol

21、, 2003. 69(8): p. 5023-8. 13. Enomoto, Y., et al., Rapid diagnosis of herpes simplex virus infection by a loop-mediated isothermal amplification method. J Clin Microbiol, 2005. 43(2): p. 951-5. 14. Ihira, M., et al., Rapid diagnosis of human herpesvirus 6 infection by a novel DNA amplification metho

22、d, loop-mediated isothermal amplification. J Clin Microbiol, 2004. 42(1): p. 140-5. 15. Yoshikawa, T., et al., Detection of human herpesvirus 7 DNA by loop-mediated isothermal amplification. J Clin Microbiol, 2004. 42(3): p. 1348-52. 16. Ihira, M., et al., Direct detection of human herpesvirus 6 DNA

23、 in serum by the loop-mediated isothermal amplification method. J Clin Virol, 2007. 39(1): p. 22-6. 17. Wakabayashi, T., et al., Rapid and sensitive diagnosis of adenoviral keratoconjunctivitis by loop-mediated isothermal amplification (LAMP) method. Curr Eye Res, 2004. 29(4-5): p. 219-24. 18. Poon,

24、 L.L., et al., Rapid detection of the severe acute respiratory syndrome (SARS) coronavirus by a loop-mediated isothermal amplification assay. Clin Chem, 2004. 50(6): p. 1050-2. 19. Hong, T.C., et al., Development and evaluation of a novel loop-mediated isothermal amplification method for rapid detec

25、tion of severe acute respiratory syndrome coronavirus.J Clin Microbiol, 2004. 42(5): p. 1956-61. 20. Poon, L.L., et al., Detection of human influenza A viruses by loop-mediated isothermal amplification. J Clin Microbiol, 2005. 43(1): p. 427-30. 21. Imai, M., et al., Development of H5-RT-LAMP (loop-m

26、ediated isothermal amplification) system for rapid diagnosis of H5 avian influenza virus infection. Vaccine, 2006. 24(44-46): p. 6679-82. 22. Ito, M., et al., Rapid detection and typing of influenza A and B by loop-mediated isothermal amplification: comparison with immunochromatography and virus iso

27、lation. J Virol Methods, 2006. 135(2): p. 272-5. 23. Ushio, M., et al., Detection of respiratory syncytial virus genome by subgroups-A, B specific reverse transcription loop-mediated isothermal amplification (RT-LAMP). J Med Virol, 2005. 77(1): p. 121-7. 24. Shirato, K., et al., Diagnosis of human r

28、espiratory syncytial virus infection using reverse transcription loop-mediated isothermal amplification. J Virol Methods, 2007. 139(1): p. 78-84. 25. Okafuji, T., et al., Rapid diagnostic method for detection of mumps virus genome by loop-mediated isothermal amplification. J Clin Microbiol, 2005. 43

29、(4): p. 1625-31. 26. Yoshida, N., et al., Simple differentiation method of mumps Hoshino vaccine strain from wild strains by reverse transcription loop-mediated isothermal amplification (RT-LAMP).Vaccine, 2007. 25(7): p. 1281-6. 27. Toriniwa, H. and T. Komiya, Rapid detection and quantification of J

30、apanese encephalitis virus by real-time reverse transcription loop-mediated isothermal amplification. Microbiol Immunol, 2006. 50(5): p. 379-87. 28. Parida, M., et al., Real-time reverse transcription loop-mediated isothermal amplification for rapid detection of West Nile virus. J Clin Microbiol, 20

31、04. 42(1): p. 257-63. 29. Fukuda, S., et al., Rapid detection of norovirus from fecal specimens by real-time reverse transcription-loop-mediated isothermal amplification assay. J Clin Microbiol, 2006. 44(4): p. 1376-81. 30. Fujino, M., et al., A simple method for the detection of measles virus genom

32、e by loop-mediated isothermal amplification (LAMP). J Med Virol, 2005. 76(3): p. 406-13. 31. Fukuta, S., et al., Development of immunocapture reverse transcription loop-mediated isothermal amplification for the detection of tomato spotted wilt virus from chrysanthemum.J Virol Methods, 2004. 121(1):

33、p. 49-55. 32. Song, T., et al., Sensitive and rapid detection of Shigella and enteroinvasive Escherichia coli by a loop-mediated isothermal amplification method. FEMS Microbiol Lett, 2005. 243(1): p. 259-63. 33. Iwamoto, T., T. Sonobe, and K. Hayashi, Loop-mediated isothermal amplification for direc

34、t detection of Mycobacterium tuberculosis complex, M. avium, and M. intracellulare in sputum samples. J Clin Microbiol, 2003. 41(6): p. 2616-22. 34. Endo, S., et al., Detection of gp43 of Paracoccidioides brasiliensis by the loop-mediated isothermal amplification (LAMP) method. FEMS Microbiol Lett, 2004. 234(1): p. 93-7. 35. Hirayama, H., et al., Rapid sexing of bovine preimplantation embryos using loop-mediated isothermal amplification. Theriogenology, 2004. 62(5): p. 887-96.