プロテオーム研究における二次元電気泳動法 及び MelanieⅡ - Bio-Rad
Contents
1. 00000000 PFGE RFLPsO 0000 eu 000000000 163 212 126 148 0 0 0 0 1 T Mitsuda K Arai S Fujita and S Yokota Journal Hospital Infection 31 123 134 1995 2 T Mitsuda T Muto M Yamada K Kobayashi M Toba Y Aihara A Ito and S Yokota Journal of Clinical Microbiology 36 652 656 1998 3 R Puohiniemi T Heiskanen A Siitonen Journal of Clinical Micro biology 35 2487 2491 1997 4 F C Tenover R D Arbeit R V Goering P A Mickelsen B E Murray D H Persing and B Swaminathan Journal of Clinical Microbiology 33 2233 2239 1995 5000000000000 000000 0000000 50 1077 1086 1995 0 02. 1 McLellan 0000 3 0 80 20 40 20mM 30mM MES 30mM MOPS 35mM HEPES 30mM EPPS 25mM 40mM CAPS 20mM CAPS gt 2 1 ARAF FRIN RRS 24 791 0 0 116 014000 0000000 5 7 1800000000 28 03 5811 6270 03 5811 6272 0222 003300000000000002 7 300009 2 045 476 0351 045 476 0350 0 305 0031 1 15 1 1 0298 52 0835 0298 52 0829 0 90532 00250000000001 14 110 00000 9 06 308 6568 FAX 06 308 3064 000000 0 1465 009300 0000000 3 121 1 0 1 052 702 2358 052 702 2812 00000 90 812 00130000000000 2 175000000 9 2 092 475 4856 092 475 48586 0 72 03 5811 6271 03 5811 6272
3. 00000000 00000000000 DO O 0 0 amp 4 000000 10 200000000 L L 210 00000000000 LIF 20 000000 0 2 210 000000000000 21000000060 000 0 Dichroic Mirror Ci Ca E E E E E C E c 0000 20000 00000 0000 0 00 0 0 00 42 5
4. 00 00000000 E 0 4000 oo00oo0000 9 0000000 0000000000000 00000000000 00 000 2000 00 00000 600 5 000 0 0 0000 0 0 00000 00000 0000 000 00000 000
5. 000000000000000000 O NaivePAGE pH D 38 10 2 000 0 0000 0 000 0 3 00 000000 Meedian 00000 0 pH9 4 1021 0 00 000 0 00 0 0 0 0 0 0 0 L 000000000 00000 00 0 0 0 0 0 0 9510 00000000000 5250 0 0 0 59910000 000 0 0 0 0 0 0 0 0 0 0 5250 0 0 0 0 0 00000
6. 2D Two dimensional polyacryl amide electrophoresis 000 000000 2010 00000 20 19801 0 O0 0 0 Immobilized pH Gra dient IPG 0 0000 O pH3 120 000000 0000 0000 0 00 00 0 0 0 0 00 00 0000000000 0 00000 00 O 16 18cm 00000000000 00 3000 00 00000 PGI 000 0 1 6 000 0 O 130 130 0000 00 0 0 0 00 00 143 3 protein kinase inhibitor 00 00000000 0000 0000 29
7. 0000 0 00 L Centers Disease Contorol Prevention wHop 000 00000 060 00 0 0 00 00 000 E e DNAD 00 00000 0 0 Eppendorfi 0 157 proteinase Eppendorf
8. 00000 0000 80 11 O00000 000000 00 0 0 0000 000000 000000000 00 0000 0 0 EEU E EA E
9. mutant wild type 99 Wild Partially melted Mutant Partially melted mutant wild type J Single _ strands gt Double strand 0O 1 0 0 DGGE 7 4 00000 0000 0 0000 00 00 0 91001000 20 701 0011 00000 000000 000000000000 0 Deen 00000000 c 0 000 0O DGGE 50 65 10000 000 0 0000 0 O 0 0 0000 0 O O GC 000000 40681 0 0 GC 000000000 DGGEL 0 0 mutant type wild type 10000 000 0000 0 0 0 0 0 0 L 0 000
10. 0000 0000000 SYBR GIO O 0 0 0 1000 100008 400 0 00 0 000 00000 0 0 0 0 00000 30 63 000000000000 25 DO DNAD 0 00 0 4 U 0 42100 00000 0 0 0 0 0 7 00000000 ili E 1 7 2 5 E 5 2 m 5 24 75u mMmIDx 37500 AAEE coated 00000 2 5 0000 0 49880 0000 1500 dsDNA Buffer 4 001 0 50 0 1406 1516 413 4176 42760 713bp 0 7266 00 0000000000000 0 0 000 A E 600 1106 1116 O 4896 5015 000 00000 0 24cm 0 5 0 6 2
11. 2D 0 0 ExPASy Expert Protein Analysis System 0 0 0000000000000 00 ch2d O0 0000000 Melanie 1991 0 0 0 0 O Ron D Appeli C 00000000000000 000 BioRad Melanie 4A000000 Meanie 0 C 0 0 0000000 MelBatch MelBatch Informations Mater gel Melanie 2 D
12. 0 0 0 0 0 000 O00000 0000 00000 00000000000000 0 29940 0 00 000 00000000000 Proteome Protein 100 00000 0000 00000 0000 0
13. Application News Bio Rad Laboratories 1998 MAY 1 4 6 E 2 e a 9 SA 11 L 0000000000 19861 1 00000 00000 0 0000 0 0 00 0000000 0 2005 0000000000000 0000000 19951 00 00000 00000000 Haemophilus 0 0000000 000 0 0 00 0 00 00000 1299 10 0 0 0 0 50 0 000000000 0 0 0 00 0 0 00 0 0 00
14. e 1 PFGE ODOL 0000000 Tips 00 1990 5 0 0 000 0 0 MRSA 00 0 00000 O golden standard ECEE B E 19961 0 0000000 11 0000000000 9157 00000000000 925 8001 0 00000 0000000000000 90 000 000 m s HE EEEO
15. 1 Fisher S And Lerman L Proc Natl Acad SDI 80 1579 1583 1986 2 Myers R Fisher S Lerman L and Maniatis T Nucl Acids Res 13 3131 3145 1985 3 Costes B Girodon E Ghanem N Chassignol M Hum Mol Genet 2 393 397 1993 4 Lerman L S SilverStein Methods Enzymol 155 482 501 1987 5 Gelfi C Righetti P Cremonesi L and Ferrari M Electrophresis 15 1506 1511 1994 6 Steger G Nucleic Acids Res 22 2760 2768 1994 00000000 00000000000 0000 2 Bio Focus LIF 0 0 0000000000000 0 000001 00000 2100000000000000 0 21210 000000000600 0 00 0 0 0 000000000600 L
16. Bulletin 2043 2216 Anderson P Heron I Simultaneous electroelution of whole SDS polyacrylamide gels for the direct cellular analysis of complex protein mixtures Journal of Immunological Methods 161 29 39 1993 Teena 0000 m 000 19980 70 300 8 BIO RAD Q amp A 65x 5 5 Lx W 16 Lx W 000000 00000 00 0 0 0 0000000000 00 Stan Copper Stain Kit A UOU E BEE ENEE UE HE E E UU 001 A U CE EE EE E E E 000 000000000060 7000000000000 0000000000 0000 00000
17. L 00000 Tumor Normal A 7 C CEE E E PAO 0 80 000 0 00 0 000 0 00 0 0 00 e ee SYPRO 100000000000 8 Bio Rad Laboratories 00000 00 00 00 00 00001 000 000000 00000000 00000000000 0000000000 00 0 E 00 0 0 0 00 00 0 0 0 0 000 0 0 0 0 0 0 0
18. 0000 numer Intemet server ExPASy etc SWISS 2DPAGE Databases Swiss Prot etc 1 000 280 00 Meanie 0 0 0 0000 00000 20 28100 0 00 0 0 0 00 0 00 0 0 0 000 Melanie 0 0 5 55 20 0 0 16 0 0 280 0000000000000 version 22 000000 0000000 00 000 00 0 00 0 0 000 0 ExPASy exspasy hcuge ch 00 0 0 0 00 0 0 00 000000000000 00 50 000 SMSSPROT O Accession 0 00000000000 210 000 00 00000 O Sun Microsystems Work stations 0000000
19. _ isphyed 1 0 of 1791 bases 4 000 000 000 000 00 68 82 0 0 00000000 5468 4100 L 0 000 DGGE 1 000000000000 30 406 O0 0 0 D E O D L cystic fibroblasti exon10 369bp 11 289bp 14 O0 0 0 O 0 00O 0O 0000000 4 00000 00 0000 4 0 O O O Dr L Silverman University of North Carolina School of Medicine O00 00000 16 16 6 Urey 6 0 O000000 00 37 5 DO 125 0 0 00 000 PCR 000000 200 300 00000 Sa 0 2x 7 Glycerol 0 05 2mM EDTA 00000000 00000 BWI 5 0 0 5060 0 Ramp Rate 2 DCodej 00000 0000000 00 1000 00010 05 10 11 1 2 3 4 5 6 7 e 9 10 11 12 13 14 5 Separation of amplified mutant and wild type alleles of exons 10 and 11from the cystic fibrosis gene on the Dcode system Lane 1 mutant allele AF508 Lane 2 wild type allele Lane mutant allele 520 Lane 4 mutant allele R560T Lane 5 9 11 13 wild type allele lane 6 exon 11 mutant allele 1717 16 lane 7 mutant allele 05515 lane 8 m
20. 0 000 0 000 0 0 0 425 0000 O O0 0O 0O L 0000000 E O O L 0000 00000000 wid type mutant 2100000 2930 0 00 0 0 2 55 0 00 0 00 0 0000000905 20 310 0 0 0 0 0 0 0 0 00000 0 00 0 O O O 16x20cm 0 75mm 0 810 0 0 0 0 0 0 0 37 51 10 0000 0 0 00000000000000 000 00 0 amp 30w 000 mutant O 0 OOU LE Bulletin 2104 00000 5 SSCP samples run at 8 C on the DCode system Lane 1 heterozygouz mutant IVS1 6 lane 2 wild type lane 3 homozygous mutant B sickle 2 000 0 0 0 0 00000000000000600 10 00 0 0 0 0 0 0 000 0000 0000 00000 000 0 0 0 55 00000 0 DGGE TTGED 0 0 0 0 000 E 000000 00000000000000 0000000 E C
21. 0 0000 wid 30 O 1 51 1 IVS16 51 00 mutant O O0 00000 4i 0 00O O DNAD 000 600 0 0 0 4065 00000 281bp 00000 0 200 7 5 10 1 0 81000 00 0 0700 37 5 0 0 0 20 700 0 0000000 0 1 310 0 5691 0000000000 41 B O EEUE E OA 00001 2 20 701 00000000000 45900 0 0 0 0 00 31 0 Heterodupiex 41000 0 00 Bulletin2107 0 0 0 000 L 1 2 3 4 5 2 Perpendicular DGGE 3 separation of separation of B thalassemia thalassemia samples mutant IVS1 6 run on the the DCode system Lane 1 DCode system The optimum compound mutant sample denaturant concentration to IVS1 1 IVA1 6 lane 2 use gel was deter mutant sample IVS1 1 lane mined at the location where mutant sample IVS1 6 lane 4 the maximum split occurs mu
22. 0 LL 00000 DNAD 000 0000 0 0 0 000 000 0000 00000 Bio Rad Laboratories 00000 00 00 0 0 0 00 000 0 000000000000 00000 000 O American Type Culture Collection Genomic 00000 000 000 00000 0 00000 1
23. 000000 0 Meanie 000000000000 00 0 version upi 00 0 00 000 NCIFlicker http wwwHech ncifcrf gov flicker 9 Melanie 000 000 0 00 00 0 0 00000 0 0 1 0000 0 0 000 0 0 0 000 1 Proteome Research New Frontiers in Functional Genomics 1997 edited by D F Hochstrasser Springer Verlag Berlin 2 HsichG et al N Engl J Med 1996 335 824 930 3 Appel RD et al Electrophoresis 1997 18 2724 2734 4 Hooglnd C et al Electrophoresis 1997 18 2755 2758 5 Lemkin P F Electrophoresis 1997 18 2759 2773 1 A 2 2 http www expasy ch ch2d 2d index htmi SWISS 2DPAGE http www expasy ch ch2d ch2d top html YPM http www ibgc u bordeaux2 fr Y P M YEAST 2D PAGE http yeast 2dpage gmm gu se ECO2DBASE http pcsf brcf med umich edu edu eco2dbase S
24. 00000000000000001 0000 00000 00000000 00000000000 1 0000 0 00 Single Strand Conformational 55 Denaturing Gradient Gel Electrophoresis Temporal Temper ature Gradient Gel Electrophoresis TTGE 1000 0 000 0 000 0 0 0 0000000000 0000 001 2 00000000000000 DGGE Gradient Electrophoresis CDGE Constant Denaturant Gel Electrophoresis 0 0 L 0000000000 000 DGGE 1001 000000000 000 0000000 000 DGGE 0000000 Fishen 000000000000 0000000000000 00000 Tmi 00000 0 00 0 00 00 000
25. 006 000000000 0 00 0 00000 1 lt 2 00000000009 c 0 2 46 000000000 00000 00 00000 00000000000000 505 0 00000 0 0 001 200 gt gt 6 0
26. tant sample IVS1 110 lane between the heterozygous 5 wild type DNA and homozygous alleles 3 00 0000000 Temporal Temperature Gradient Gel Electrophoresis 000 0 O0 DGGELU 0 0 0 DNAD 0 000 Mutant Wild 00000000000000000000 Ramp 00000000000 00000 000 000 0 00 000 DNAD 00 0 26810 0000 00000 066 Bio Rad Laboratories 40206 1 E 20 0 000000 0000000060 0000001 Ramp 000 66000 00 0 0 0 0 OOOD A
27. ub2D http pc13mi biologie unipgreifswald de Cyano2Dbase http www kazusa or jp tech sazuka cyano proteome html Aberdeen 2 D db http www abdn ac uk mmb023 2dhome htm Maise 2 D db http moulon moulon inra fr imgd Fly 2 D db http tyr cmb ki se LSB 2 D db http www Isbc cony2dmaps patterns htm HSC 2DPAGE http www harefield nthames nhs uk nhli protein HEART 2DPAGE http www chemie fu berin de user pleiss dhzb htmi HP 2DPAGE http www mdc berin de emu heart Danish 2 D db http www biobase dk cgi bin celis Emb stem cell http www ed ac uk nh 2DPAGE A375 UCSF 2 D http rafael ucsf edu 2DP AGE home htmi ANGL Breast 2 D http www anl gov CMB P MG projects index_hbreast html Colon carcinoma http www ludwig edu au www jpsl jpslhhome html 000 pp 127Tabie5 1 000 018 SWISS 2DPAGE map version 2 2 Human biological map Colorectal epithelia cells Cerebrospinal F luid Erythroleukemia Cell H epG2 HepG2 secreted proteins Human promyelocytic leukemia cells Liver Kidney Lymphoma 10 Plasma 11 Platelet 12 Red blood cells 13 Macrophage like cell line Others 14 Dictyostelium discoideum 15 E coli 16 Yeast Sanchez etal 1995 Electrophoresis 16 1131 1151 Appel et al 1996 Nucleic Acids Res 24 180 181 paR al me
28. utant allele G542X lane 10 mutant allele R553X lane 12 mutant allele 05510 lane 14 mutant allele 5549 Ramp Rate Mac M metj 1 141000000 2100000000 2050 91 10 13 wildtype 0 0 0 00000000000000 000000 00000000 2103 000 000 4 000 55 Single Strand Conformational Polymorphisms wid type mutant 0 0 0 0 0 DNAD OLDU 55 E E E E Eee E EE 000 0000 0 0000 0 00 0 0 0 0 00 5 60 00000000 0000000 1 0000 5 00
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