Target Discovery and Validation Reviews and Protocols

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213 K6a K6a Hair follicles; Lysis of the nail bed Pachyonychia 97 (see Fig. 3E) K6b K6b activated epithelium; neonatal congenita-like K17 K17 keratinocytes death around P3 nail lesions K7 K7 Unknown K8 K8 Simple a) Embryonal death Ulcerative 53–55 epithelia ED12.5; trophoblast colitis giant cell fragility (C57BL/6x129X1SvJ) b) Colorectal hyperplasia, colitis, rectal prolapse, predisposition for liver injury (FVB/N) n.d; not done. a Orthologous human and mouse keratin genes of each clusters I and II are compared in a separate table Tables 1 and 2 follow the new and open nomenclature based on the principles of the HUGO Gene Nomenclature Committee. A comparative table aligning the old nomenclature to the novel system is available as a guideline in Magin et al. (4). The major expression and knockout phenotype is briefly outlined in all cases where keratin-deficient mice are generated and characterized. Double- and triple-keratin knock outs are shaded in gray. The human pathology that could be deduced from a mutated keratin gene is briefly outlined but does not inevitably coincide with the phenotype of mice with a targeted deletion in the orthologuous keratin gene. (C57BL/6x129X1SvJ) and (FVB/N) are mice strains.
214 Vijayaraj, Söhl, and Magin ulcerative colitis and Crohn’s disease (51,52). Crohn’s disease and ulcerative colitis, together known as inflammatory bowel disease, are chronic illnesses of unknown origin. The inflammation within the intestinal tract (within the colon in ulcerative colitis and anywhere from the mouth to the anus in Crohn’s disease) leads to some or all of the following clinical symptoms: diarrhea (with or without blood), abdominal pain, fever, and fatigue. 1.1.5. Phenotypes of Keratin Knockout and Transgenic Mice 1.1.5.1. SIMPLE AND EMBRYONIC EPITHELIA Ablation of keratins expressed in simple epithelia and during embryogenesis only partially helped to identify their functional significance. Side effects because of the strain background apparently influence the phenotypical outcome of generated knockouts. Keratin K8-deficient mice, initially thought to die from hepatocyte fragility leading to fetal liver bleeding at ED12.5 in the C57BL/6 strain (53,54), have been later demonstrated to collapse from placental malfunctions (55). However, keratin K8 deficiency crossed into FVB/N strain results in viable offspring featuring colonic hyperplasia, diarrhea, and colitis (53,54). Mistargeting of apical H + /K + -ATPase-β, anion exchanger AE1/2 and the Na + transporter might lead to the observed phenotype, possibly because of the net Na + absorption associated with the Cl – secretion in jejunal epithelia (33,56). The colonic hyperplasia and colitis were later determined as chronic spontaneous helper T-cell 2 colitis that probably arises from colonic bacterial infection (57). Either distinct modifier genes or the second type II keratin, K7, expressed early during development, might compensate for the loss of K8 in FVB/N K8(–/–) mice (Tables 1 and 2). Hepatocytes from K8(–/–) mice are more susceptible to mechanical stress and hepatotoxic drugs and secrete less bile acids and ecto-ATPase, a phenomenon that is mechanistically still unexplored (58). Concanavalin A (ConA)-induced liver damage left K8(–/–) [and also K18(–/–)] epithelial cells approx 100 times more sensitive to TNF-induced cell death and increased both downstream targeted JNK- and nuclear factor-κB-protein levels. ConA-induced immunoresponse through activated T-cells was therefore thought to release TNFα, acting on the TNF receptor (TNFR)2 (59). Moreover, glutathione S-transferase-fusion protein binding assays suggested an interaction of the N-terminal parts of K8 and K18 with the TNFR2 (59). Primary hepatocytes from K8(–/–) mice being more sensitive to Fas-mediated apoptosis fall into the same category (60). Although transported by microtubules, Fas receptor abundance at the cell surface was increased in K8(–/–) mice, thus implying K8 to contribute to Fas receptor distribution (60). Concerning other keratins, a direct interaction between the TNFR1-associated death domain protein (TRADD) and the coil 1a domain of K18 and K14 was
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METHODS IN MOLECULAR BIOLOGY 360 T
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M E T H O D S I N M O L E C U L A R
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© 2007 Humana Press Inc. 999 River
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vi Preface get. Approaches to targe
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viii Contents 10 Transgenic Animal
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x Contributors SAHOHIME MATSUMOTO
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xii Contents of Volume 2 12 Validat
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2 Sioud disease pathogenesis and pe
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4 Sioud A more fruitful approach fo
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6 Sioud both transient and permanen
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8 Sioud serum biomarkers. This syne
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10 Sioud Fig. 2. Schematic of targe
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12 Sioud 13. Magin, T. M. (1998) Le
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Harnessing the Human Genome 15 The
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Harnessing the Human Genome 17 Fig.
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Harnessing the Human Genome 23 The
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Harnessing the Human Genome 27 repr
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Harnessing the Human Genome 29 14.
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Harnessing the Human Genome 31 45.
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34 Imoto et al. (4,5), and Bayesian
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36 Imoto et al. 2. Materials Two ty
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38 Imoto et al. Fig. 3. Graphical v
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40 Imoto et al. Fig. 5. Result of t
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42 Imoto et al. p(D |G) = ∫ p(D,
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44 Imoto et al. β k (k = 1,…,q j
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46 Imoto et al. Fig. 7. Partial res
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48 Imoto et al. Fig. 8. Strategy to
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50 Imoto et al. Fig. 9. (continued)
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52 Imoto et al. Table 3 List of Roo
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54 Imoto et al. 5. De Hoon, M. J. L
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56 Imoto et al. 39. Kamimura, T., S
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58 Beaty et al. cytotoxic drugs and
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60 Beaty et al. Fig. 1. Principles
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62 Beaty et al. oligonucleotide mic
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64 Beaty et al. 3. The 12% polyacry
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66 Beaty et al. 2.3.3. Protein Stai
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68 Beaty et al. 3.1.3. Labeling of
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70 Beaty et al. 1 µL of the anneal
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72 Beaty et al. 4. Phenol:cholorfor
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74 Beaty et al. 19. Wash twice with
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76 Beaty et al. The Following Volum
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78 Beaty et al. using a protein ass
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80 Beaty et al. 6. Shake the gel fo
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82 Beaty et al. Fig. 2. Preparation
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84 Beaty et al. search in. A widely
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86 Beaty et al. 3. Kern, S. E., Hru
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88 Beaty et al. 34. Peters, D. G.,
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5 Molecular Classification of Breas
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Molecular Profiling of Breast Cance
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Fig. 2. (Continued) the right ident
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6 Discovery of Differentially Expre
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Gene Target Discovery 117 In the fi
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Gene Target Discovery 119 sequences
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Gene Target Discovery 121 There is
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Gene Target Discovery 123 digestion
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Gene Target Discovery 125 Fig. 1. P
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Gene Target Discovery 127 5. Sargen
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Gene Target Discovery 129 41. Berry
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132 Matsumoto, Miyagishi, and Taira
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144 Fig. 1. The ribozyme-expression
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146 Sano and Taira 5. 10X L (low-sa
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148 Sano and Taira ribozymes may cl
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150 Fig. 3. A system for the identi
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152 Sano and Taira 4. Notes 1. We r
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9 Production of siRNA- and cDNA-Tra
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Transfected Cell Arrays 157 Fig. 1.
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Transfected Cell Arrays 159 2. The
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Transfected Cell Arrays 161 5. Simp
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264 Skovseth, Küchler, and Haralds
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270 Iversen and Sørensen witnessed
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272 Iversen and Sørensen Fig. 1. P
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274 Iversen and Sørensen the core
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14 An Overview of the Immune System
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Overview of Immune System 279 diffe
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Overview of Immune System 281 Fig.
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Overview of Immune System 283 then
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Overview of Immune System 285 expre
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Overview of Immune System 289 solub
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Overview of Immune System 291 amino
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Overview of Immune System 293 (unre
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Overview of Immune System 297 the c
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Overview of Immune System 299 (44).
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Overview of Immune System 301 demon
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Overview of Immune System 307 some
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Overview of Immune System 309 sera
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Overview of Immune System 313 measu
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Overview of Immune System 315 42. H
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Overview of Immune System 317 74. D
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15 Potential Target Antigens for Im
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Tumor Antigen Discovery 321 develop
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Tumor Antigen Discovery 323 panel b
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Tumor Antigen Discovery 325 16. Joc
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16 Identification of Tumor Antigens
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Immunoproteomics 329 by “shot gun
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Immunoproteomics 331 isoelectric fo
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Immunoproteomics 333 3. 2D-PAGE is
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17 Protein Arrays A Versatile Toolb
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Protein Arrays 337 Table 1 Classifi
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Protein Arrays 339 fractions from c
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Protein Arrays 341 combinatorial sc
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Protein Arrays 343 The ideal proteo
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Protein Arrays 345 18. Cekaite, H.
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Protein Arrays 347 49. Yuko Kawahas
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Index 349 Index A Acetyl-CoA carbox
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Index 351 conditional by using FRT
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Index 353 Recombinant tumor antigen
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