The Genom of Homo sapiens.pdf

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COMPARATIVE GENOMIC ANALYSIS 269µg/ml E. coli genomic DNA, 0.2% Tween 20, and 10 mMcitric acid, pH 6.0, prepared as described at http://zfin.org/zf_info/zfbook/chapt9/9.82.html) with 48 and 72hours postfertilization (hpf) zebrafish embryos in a 96-well microtiter plate (VWR Scientific, cat. #62402-933),stained for 4 or 8 hours to develop the characteristic darkcolor, washed, and photographed by a CCD-cameraequippedLeica MZFLIII fluorescence stereo-microscope.These results clearly indicate that single-strandedDNA probes ranging in size from 1 kb to 100 bases canbe used for successful in situ hybridizations to zebrafishembryos in a 96-well format.This robust protocol for zebrafish embryo wholemountin situ hybridization using single-stranded DNAprobes has allowed us to multiplex our experiments efficientlyand thus begin to investigate several additionalpredicted human Chromosome 22 genes based on thisexon-specific PCR, unidirectional amplification approach.As another example of our optimized protocols, we investigatedthe expression of a human Chromosome 22gene which has an unknown function, but whose expressionhas been observed through EST studies. This predictedgene corresponds to an EST, KIAA0819 (Nagaseet al. 1998), and the results of our analysis are shown inFigure 4. Hybridization of a single-stranded unidirectionalDNA probe (ssDNA) corresponding to exon 1 inone copy of the dual-copy zebrafish ortholog of mouseKIAA0819, as revealed by the PIP, is shown in Figure 4a.The hybridization results presented in Figure 4b indicatethat only the antisense probe gives a positive hybridizationsignal to the otic vesicle at 24, 48, and 120 hpf.In contrast to the above results, in the two experimentsdescribed below we have observed hybridization not onlywith the antisense probes corresponding to the respectivemRNAs, but also with the sense probes corresponding tothe antisense RNAs for two genes in the zebrafish systemthat correspond to two predicted human Chromosome 22genes AP000553.6 and AP000354.2, as shown in Figures5 and 6, respectively.Human AP000553.6Ensembl GeneID ENSG00000161179, located on humanChromosome 22 from 20306935 to 20308868 bp, isa novel predicted gene that has no known function or expressiondata. This unknown gene contains five predictedexons in humans and its orthologs in mouse, rat, fugu, andzebrafish that were found by a reciprocal blast search.The PIP shown in Figure 5a indicates that this gene hashighly conserved exonic sequences in mammals butshows a lower level of sequence conservation in fish,where several exons are less than 50% conserved.To study the expression profile in zebrafish, exon-specificprimers were selected to PCR-amplify the exon regionfrom zebrafish genomic DNA. For the expressiondata shown in Figure 5b–d, the 337-bp 3´-most exon(exon 5, Ensembl ExonID ENSDARE0000013070) wasamplified using a sense primer 5´TGAGCACCATGGGTAAGAAC3´ beginning at the 24th base of the exon andan antisense primer 5´GGATCCCTTGTTGCCTGTAA3´ beginning at the 317th bp of the exon. Theseprimers generated a 293-bp PCR product that then wasused to generate ssDNA sense and antisense probes in thepresence of the DIG-labeled dNTP mix.As shown in Figure 5b, no background hybridizationwas observed in the absence of probes when the antisenseand sense probes shown in Figure 5c and d were incubatedin parallel. As indicated in Figure 5c, the antisensehybridization probe reveals the presence of hybridizingRNA at 12 hpf in the mesoderm layer, at 24 hpf in notochord,at 48 hpf in both notochord and otic vesicle, and at72 hpf in the otic vesicle and liver. In contrast, Figure 5dshows that the ssDNA sense probe hybridizes to RNA inthe notochord at 24 hpf through 72 hpf, indicating thelikely presence of an antisense regulatory RNA for thisgene product expressed in the notochord that may play arole in suppressing the production of the AP00553.6 proteinat the translational level.Human AP000354.2Ensembl GeneID ENSG00000100014, located onChromosome 22 at positions 23041871–23137773 bp, isa novel predicted gene that has no known function or expressiondata. This unknown gene contains 13 predictedexons, and a reciprocal blast search revealed orthologs inmouse, rat, fugu, and zebrafish. The PIP shown in Figure6a reveals the regions of sequence similarity that are 50%or greater, and, as described above for the AP000553.6gene, the exons of this AP000354.2 gene also are highlyconserved in mammals but less conserved in fugu and zebrafish.Interestingly, of the 12 exons in mammals, only5 are conserved recognizably in zebrafish, with 4 beingmore than 50% conserved.To investigate the expression profile of the AP-000354.2 gene, PCR primers were picked to amplify thesecond, 331-bp zebrafish orthologous exon. Here thesense primer, 5´CAAACGGACAACATCCATGA3´,was 11 bp into the exon, and the antisense primer was5’CCTCATGTCCCTCAGCTCAC3´. These two primersgenerated a 300-bp PCR product that served as a templatefor the unidirectional PCR to generate sense and antisenseprobes.As shown in Figure 6b, although there is not much differencebetween the antisense and sense probe hybridizationin the 12 hpf embryos, in the 24 hpf embryos, hybridizationin the telencephalon portion of the forebrain,the midbrain, and the hindbrain is not visible with thesense probe, whereas the notochord and myotomes showstrong hybridization with both antisense and senseprobes. In the 48 hpf embryos, the myotomes and telencephalonportion of the forebrain seem to hybridize morestrongly with the sense probe. In the 72 hpf embryos, thetelencephalon portion of forebrain is more intensely hybridizedwith the sense probe, whereas the lateral lineganglia show similar hybridization to both the antisenseand sense probes. Other features, including the disappearanceof staining in notochord and myotomes, aresimilar with both probes. This complex expression pattern,observed for both antisense and sense probes, indicatesthat the AP000354.2 gene is expressed in multiple
270 ROE ET AL.aNovel gene KIAA0819[AB020626]11 9ratmousefugu1fugu2zeb1zeb2200k 202k 204k 206k 208k 210k 212k 214k 216k 218k 210kNovel gene KIAA0819[AB020626]8 7 6ratmousefugu1fugu2zeb1zeb2220k 222k 224k 226k 228k 230k 232k 234k 236k 238k 240kratmousefugu1fugu2zeb1zeb2Novel gene KIAA0819[AB020626]5 4 3 2 1100%50%240k 242k 244k 246k 248k 250k 252k 254k 256k 258k 260kb anti-sense kiaa0891 sense kiaa0891 probe no probe control120 hpf48 hpf24 hpfFigure 4. (a) MultiPIP analysis of the predicted genes from human, rat, mouse, fugu, and zebrafish with homology to cDNA probeKIAA0819. The unique unidirectional 708-base ssDNA probes from exon 1 of zebrafish 1 were DIG-labeled after PCR amplificationof the 708-bp region of zebrafish genomic DNA indicated by the red rectangle. This probe is only a 74-base region with a percentidentity of 79.7 out of 708 bases in zeb2 with an overall identity of less than 10%. (b) Whole-mount in situ hybridization of thessDNA antisense, ssDNA sense, and no-probe control described in Fig. 7 to 24 hpf, 48 hpf, and 120 hpf zebrafish embryos indicatinghybridization to the otic placode region only with the DIG-labeled antisense probe.
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ForewordIn 2001, as we considered t
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The Finished Genome Sequence of Hom
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4 ROGERSFigure 2. Accumulation of h
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6 ROGERSFigure 4. Sequencing center
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8 ROGERSabcFigure 5. Ensembl view o
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10 ROGERS2000. Analysis of vertebra
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The Human Genome: Genes, Pseudogene
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VARIATION ON CHROMOSOME 7 15rived f
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VARIATION ON CHROMOSOME 7 17DNAs an
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VARIATION ON CHROMOSOME 7 19expecte
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VARIATION ON CHROMOSOME 7 21Drosoph
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Mutational Profiling in the Human G
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HUMAN MUTATIONAL PROFILING 25Anothe
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HUMAN MUTATIONAL PROFILING 27Figure
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HUMAN MUTATIONAL PROFILING 29Rieder
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32 SCHMUTZ ET AL.algorithm itself,
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34 SCHMUTZ ET AL.Figure 2. Genomic
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36 SCHMUTZ ET AL.compared. Some of
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Human Subtelomeric DNAH. RIETHMAN,
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HUMAN SUBTELOMERIC SEQUENCES 41The
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HUMAN SUBTELOMERIC SEQUENCES 43cate
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HUMAN SUBTELOMERIC SEQUENCES 45Figu
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HUMAN SUBTELOMERIC SEQUENCES 47pres
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50 COLLINSand expand the genomics r
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52 COLLINSFigure 2. A public-sector
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54 COLLINSdefine all the parts of t
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56 BENTLEYmon over many generations
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58 BENTLEYTable 1. Genetic Disease
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60 BENTLEY(Clark et al. 1998; Reich
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62 BENTLEYACKNOWLEDGMENTSThe author
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SNP Genotyping and Molecular Haplot
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GENETIC ANALYSIS OF DNA POOLS 67gen
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70 FAN ET AL.matrix is then mated t
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72 FAN ET AL.Figure 3. Views of gen
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74 FAN ET AL.including 32 duplicate
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76 FAN ET AL.Figure 7. Allele-speci
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78 FAN ET AL.microsphere-based assa
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80 BERTRANPETIT ET AL.function, may
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82 BERTRANPETIT ET AL.diversity in
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84 BERTRANPETIT ET AL.gree of block
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86 BERTRANPETIT ET AL.Figure 1. Dec
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88 BERTRANPETIT ET AL.1999. Populat
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90 WINDEMUTH ET AL.Expression data.
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92 WINDEMUTH ET AL.Table 1. A Summa
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94 WINDEMUTH ET AL.Table 2. Signifi
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96 WINDEMUTH ET AL.Table 3. Summary
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98 WINDEMUTH ET AL.Table 6. List of
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100 WINDEMUTH ET AL.Table 6. (Conti
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102 WINDEMUTH ET AL.Table 6. (Conti
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104 WINDEMUTH ET AL.much of a surpr
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106 WINDEMUTH ET AL.Given our resul
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Genetic Variation and the Control o
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GENETIC CONTROL OF TRANSCRIPTION 11
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GENETIC CONTROL OF TRANSCRIPTION 11
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Genome-wide Detection and Analysis
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RECENT SEGMENTAL DUPLICATIONS 117Fi
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RECENT SEGMENTAL DUPLICATIONS 119St
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RECENT SEGMENTAL DUPLICATIONS 121Co
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RECENT SEGMENTAL DUPLICATIONS 123Ho
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The Effects of Evolutionary Distanc
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EVOLUTIONARY DISTANCE AND GENE PRED
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EVOLUTIONARY DISTANCE AND GENE PRED
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Lineage-specific Expansion of KRAB
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EVOLUTION OF ZNF GENES 133Figure 2.
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EVOLUTION OF ZNF GENES 135Figure 4.
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EVOLUTION OF ZNF GENES 137get gene,
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EVOLUTION OF ZNF GENES 139Y., Goodw
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Sequence Organization and Functiona
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CENTROMERE ANNOTATION 143THE CENTRO
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CENTROMERE ANNOTATION 145Figure 4.
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CENTROMERE ANNOTATION 147CONCLUSION
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CENTROMERE ANNOTATION 149Schueler M
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152 PARKHILL AND THOMSONFigure 1. T
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154 PARKHILL AND THOMSONshow very h
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156 PARKHILL AND THOMSONGene Loss a
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158 PARKHILL AND THOMSONYersinia ad
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160 MCKAY ET AL.Choosing Candidate
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162 MCKAY ET AL.new comparative too
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164 MCKAY ET AL.rich. Based on a th
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166 MCKAY ET AL.Embryonic Muscle an
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168 MCKAY ET AL.native polyadenylat
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Building Comparative Maps Using 1.5
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HUMAN CHROMOSOME 1p IN THE DOG 1731
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HUMAN CHROMOSOME 1p IN THE DOG 175(
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HUMAN CHROMOSOME 1p IN THE DOG 177l
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180 GEORGES AND ANDERSSON5. There i
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182 GEORGES AND ANDERSSONplied to r
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184 GEORGES AND ANDERSSONbe common
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186 GEORGES AND ANDERSSONin humans
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Evolving Methods for the Assembly o
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ASSEMBLING LARGE GENOMES 191Figure
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ASSEMBLING LARGE GENOMES 193tant ad
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Mouse Genome Encyclopedia ProjectY.
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MOUSE GENOME ENCYCLOPEDIA PROJECT 1
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DNA Sequence Assembly and Multiple
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EULERIAN ASSEMBLY AND MULTIPLE ALIG
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Ensembl: A Genome InfrastructureE.
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ENSEMBL 215projects often submit th
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Page 231 and 232: 224 ZHANGWe are waiting for experim
Page 234 and 235: Ontologies for Biologists: A Commun
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Page 245 and 246: 238 JOSHI-TOPE ET AL.Figure 1. The
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Page 252 and 253: The Share of Human Genomic DNA unde
Page 254 and 255: DNA UNDER SELECTION FROM HUMAN-MOUS
Page 262 and 263: Detecting Highly Conserved Regions
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Page 273 and 274: 266 ROE ET AL.noncoding regions. On
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Page 295 and 296: 288 OVCHARENKO AND LOOTSsequencing
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Page 318: High-Throughput Mouse Knockouts Pro
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100 bp ladder68G1168G1168H1168H6100
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FUNCTIONAL ELEMENTS IN HUMAN DNA 32
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High-resolution Human Genome Scanni
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HUMAN GENOME SCANNING 325false-posi
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HUMAN GENOME SCANNING 327affecting
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HUMAN GENOME SCANNING 329methods fo
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332 MALEK ET AL.Figure 1. The bacte
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334 MALEK ET AL.J., Vincent S., and
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336 HARDISON ET AL.reflect blocks o
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338 HARDISON ET AL.plain the region
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340 HARDISON ET AL.CALIBRATION OF T
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342 HARDISON ET AL.PositionRP2.3noE
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344 HARDISON ET AL.cific chromosoma
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346 WESTON ET AL.these differences
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348 WESTON ET AL.els controlled by
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350 WESTON ET AL.ures prominently i
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352 WESTON ET AL.nal and Bop, which
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354 WESTON ET AL.ablp 1466 bopbcrtB
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356 WESTON ET AL.like fold (Fig. 6)
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Implications of Genomics for Public
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GENETIC EPIDEMIOLOGY 361lytic epide
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GENETIC EPIDEMIOLOGY 363curate risk
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A Model System for Identifying Gene
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PTC TASTE GENETICS 367Figure 2. Hap
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PTC TASTE GENETICS 369Table 2. Hapl
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PTC TASTE GENETICS 371the emergence
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374 MCCALLION ET AL.Figure 1. Schem
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376 MCCALLION ET AL.lier (Carrasqui
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378 MCCALLION ET AL.Table 3. HSCR A
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380 MCCALLION ET AL.Figure 3. Trans
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Genetics of Schizophrenia and Bipol
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SCHIZOPHRENIA AND BIPOLAR AFFECTIVE
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The Genetics of Common Diseases: 10
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GENETICS OF COMMON DISEASES 397with
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GENETICS OF COMMON DISEASES 399SELE
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GENETICS OF COMMON DISEASES 401F.,
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404 CHEUNG ET AL.netic analysis. Ex
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406 CHEUNG ET AL.Figure 3. The expr
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Regulation of α-Synuclein Expressi
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α-SYNUCLEIN EXPRESSION AND PD 411T
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1. The levels of α-synuclein prote
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α-SYNUCLEIN EXPRESSION AND PD 415g
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418 BOTSTEINFigure 1. (A) Blectron
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420 BOTSTEINFigure 3. Cluster diagr
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422 BOTSTEINFigure 6. Kaplan-Meier
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424 BOTSTEINGarber M.E., Troyanskay
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426 ANTONARAKIS ET AL.1316192225283
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428 ANTONARAKIS ET AL.Figure 5. Sam
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430 ANTONARAKIS ET AL.POPULATION VA
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432 JORGENSEN ET AL.tive small mole
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434 JORGENSEN ET AL.FLAG-tagged pro
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436 JORGENSEN ET AL.visualization t
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438 JORGENSEN ET AL.AArp2/3 Complex
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Pathway40S440 JORGENSEN ET AL.ANutr
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442 JORGENSEN ET AL.Giaever G., Chu
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Genomic Disorders: Genome Architect
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GENOME ARCHITECTURE AND GENOMIC DIS
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Human Versus Chimpanzee Chromosome-
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HUMAN VS. CHIMP CHROMOSOME COMPARIS
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HUMAN VS. CHIMP CHROMOSOME COMPARIS
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Novel Transcriptional Units and Unc
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TRANSCRIPTIONAL UNITS AND GENE PAIR
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mtDNA Variation, Climatic Adaptatio
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mtDNA VARIATION 473Figure 3. Region
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ANALYSIS OF ADAPTIVE SELECTION FORR
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mtDNA VARIATION 477Figure 8. Temper
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Positive Selection in the Human Gen
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HUMAN-SPECIFIC EVOLUTIONARY CHANGES
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488 UNDERHILLorigin episodes, each
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490 UNDERHILLhaplogroups C through
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492 UNDERHILLO (Fig. 2e) that share
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The New Quantitative BiologyM.V. OL
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NEW QUANTITATIVE BIOLOGY 497alone.
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NEW QUANTITATIVE BIOLOGY 499There w
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NEW QUANTITATIVE BIOLOGY 501ceded,
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