Index 457 Heterozygous, 447g Hierarchical shotgun sequencing, 192–196 High-throughput DNA sequencing, 184–188,197–202 Hirudin, 251 His-tag, 274–275 HIV see human imunodefiency virus HIV-1 regulation of gene expression, 333–334 tat, 333–334 see also human imunodefiency virus Homoduplex, 447g Homologous, 447g Homologous recombination, 448g Homology, 220, 448g Homozygous, 448g Horizontal transfer pathogenicity islands, 27 plasmids, 26 Human genome, 23 Human Genome Project, 86, 418, 448g Human growth hormone, 251 Human immunodeficiency virus, 309, 438 Huntington’s disease, 30, 424, 425 Hybridization, 448g colony, 127, 129, 131 dot blot, 125, 427, 429 in diagnostic tests, 423 northern, 326–328, 354 plaque, 127 Southern, 125, 427 zooblot, 125, 167, 168 Hydrophobic, 448g Hydroxyl radical footprinting, 345 I Ideogram, 29 band numbering, 30 In vitro, 448g In vitro fertilisation, 142, 426 In vivo, 448g Inclusion bodies, 261–262 Independent assortment, 159 Inducer, 448g Insertional inactivation of antibiotic resistance genes, 53–54 of the lacZ gene, 54–56 Insulin, 14, 126, 250, 251, 418 Interferon, 251 Interspersed transposon-derived repeats, 13 Intron, 11, 102–103, 210, 216, 448g Intron-exon boundary, 215 Inverse PCR, 148 Inverted repeats, 143 Isoschizomer, 51, 448g IVF see in vitro fertilization K Karyotype, 28, 29, 448g Kilobase, 448g Klenow fragment, 65, 181 Knock-out, 448g L Labeling DNA probes, 127, 130–131 end labelling, 320–321 flourescent labels, 129 labels detected immunologically, 128 nick translation, 130 radioactive labels, 128 random prime labelling, 130 lacZ, 55, 58, 96, 295–297 blue-white selection, 55, 58, 96 gene fusion studies, 295 insertional inactivation, 55–56, 58, 96 promoter probe, 337 translational fusion with, 295–297 Lambda, 448g see also bacteriophage λ Library see DNA library LINE see long interspersed elements Linkage, 159, 163, 448g Linker, 106, 448g Liposome-mediated transfer (lipofection), 272, 339 Locus, 448g Log phase, 448g Long interspersed elements, 15 LTR retrotransposon, 13 M M13 see bacteriophage M13 Macro-array see DNA array MALDI-TOF MS see matrix-assisted laser desorption/ionization time-of-flight mass spectrometry Map based cloning, 159–160, 448g Marker rescue, 448g see also complementation Mass spectrometry, 358–361 Matrix-assisted laser desorption/ionization timeof-flight mass spectrometry, 358–359 MC1R, 416 Melting temperature (Tm), 66–67, 448g Membrane proteins, 293 mapping, 293–297 Mendel, Gregor, 159 Mendelian, 449g Mendelian inheritance, 142, 384, 417
458 <strong>Gene</strong> <strong>Cloning</strong> Metallothionein, 381, 400–401 Methioine sulphoxamine (MSX), 272 Methotrexate (MTX), 272 Microarray see DNA array Micro-injection, 449g Micro-satellite DNA, 13, 412 Mini-satellite DNA, 13 Minisequencing, 420 MLPA see Multiplex ligation-dependent probe amplification Model organism, 20 Arabidopsis thaliana, 20 Caenorhabditis elegans, 20 Chimpanzee, 20 Danio rerio, 20 Drosophila melanogaster, 20 Escherichia coli, 20 Mus musculus, 20 Rattus norvegicus, 20 Sacchromyces cerevisiae, 20 Molecular beacon, 429–431, 437, 438 Molecular chaperones, 262 Molecular clock, 240 Molecular diagnostics bacterial disease, 437–438 cancer, 439–441 inherited disease, 422–436 viral disease, 438–439 Moloney murine leakemia retrovirus, 385 Monosomy, 30 Motif, 234–237, 449g MS/MS see tandem mass spectrometry Multiple cloning site, 49, 449g Multiple displacement amplification, 436 Multiple sequence alignment, 233–235, 239 Multiplex ligation-dependent probe amplification, 427, 428 Multiplex PCR, 414, 420, 431, 449g Mus musculus, 20 Muscular dystrophy, 424 Mutagen, 449g Mutagenesis, 449g in determining whether an ORF is functional, 281 signature-tagged, 152–156 site-directed, 304–309 using T-DNA, 405–406 Mutation scanning, 432–435 Myogenin, 400 N N terminus, 449g N50 length, 196 NAT see nucleic acid tests National DNA Database, 416 NCBI see National Center for Biotechnology Information National Center for Biotechnology Information, 228 NDNAD see National DNA Database N-glycosylation, 265, 268 nirB, 337, 346, 349 Non-coding DNA, 10, 16 impact on genome analysis, 16 Non-homologous recombination, 284, 381–383, 449g Nonsense suppression use in chromsome jumping, 166 Northern analysis, 326–328, 354 NotI use in chromsome jumping, 164 Nuclear run-on, 330–334 Nuclear transfer, 401–403 Nuclease protection, 319–323 Nucleic acid tests, 438 Nucleosomes, 340–341 Numerical chromosomal abnormality, 30 O Oligo (dT)-anchor primer, 324–325 Oligonucleotide, 449g allele-specific, 427 calculating melting temperature, 67 degenerate, 126–127, 132 linkers and adaptors, 106 molecular beacon, 429–431, 437, 438 primers for PCR, 66–68 primers for primer extension, 324–325 primers for sequencing, 175–179 probes, 124–127 TaqMan®, 431– 432, 438 Oncogene, 371–372, 449g Oncomouse, 371–372, 449g One-hybrid screens, 351–353 Open reading frame, 208–214, 213, 279–280, 449g Operon, 449g ORF see open reading frame Origin of replication, 36–37 Orthologue, 231, 233, 449g P Paralogue, 231, 449g Partial digest, 90, 449g PCR, 449g see also polymerase chain reaction Periplasm, 449g Pfam, 225 Phage see bacteriophage Pharmacogenomics, 449g Phenotype, 419, 449g
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Gene Cloning
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vi Contents 3.14 Designing PCR Prim
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viii Contents 8.3 Identifying Eukar
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Genome Organization 25 kb 0 2 4 6 8
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Key Tools for Gene Cloning 41 (a) (
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Key Tools for Gene Cloning 63 (a) C
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Key Tools for Gene Cloning 69 (a) 5
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86 Gene Cloning you extract DNA fro
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88 Gene Cloning Box 4.1 Preparation
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94 Gene Cloning (a) (b) Bacterial s
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96 Gene Cloning (a) Insertion vecto
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100 Gene Cloning BamHI Ap r Tet r p
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102 Gene Cloning parB Cm r parA rep
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104 Gene Cloning (a) TTTTT AAAAA TT
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106 Gene Cloning Box 4.3 Converting
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108 Gene Cloning of your gene is. A
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110 Gene Cloning A4.2. Extract chro
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112 Gene Cloning A4.8. See Section
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114 Gene Cloning Q4.15. What are th
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116 Gene Cloning dystrophin gene is
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118 Gene Cloning it is still a form
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120 Gene Cloning One obvious proble
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122 Gene Cloning The main difficult
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124 Gene Cloning (a) Labeled single
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126 Gene Cloning N terminus Phe Val
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128 Gene Cloning Box 5.3 Types of D
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130 Gene Cloning Box 5.4 Methods fo
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132 Gene Cloning the DNA probe. Bot
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134 Gene Cloning a b c d e f g h 1
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136 Gene Cloning S. cerevisiae, and
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138 Gene Cloning Q5.7. Would you ex
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140 Gene Cloning use your genomic c
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142 Gene Cloning diseases, where th
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144 Gene Cloning (a) Replicative Ta
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148 Gene Cloning DNA fragment in th
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150 Gene Cloning Bacteria transform
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152 Gene Cloning Wild-type gene Mut
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154 Gene Cloning molecule using PCR
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156 Gene Cloning digesting chromoso
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158 Gene Cloning + + Ac Avr Avr cf-
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160 Gene Cloning which are very clo
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162 Gene Cloning Box 6.3 Restrictio
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164 Gene Cloning Initial region clo
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168 Gene Cloning 6.10 Cloning of th
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172 Gene Cloning Isolation of the t
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174 Gene Cloning Two approaches wer
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176 Gene Cloning synthesize DNA de
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178 Gene Cloning Box 7.1 Denaturing
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180 Gene Cloning G A T C Figure 7.4
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182 Gene Cloning although other con
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184 Gene Cloning A large DNA fragme
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186 Gene Cloning chain reaction, fl
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188 Gene Cloning rounds of amplific
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190 Gene Cloning Contig 2 Clone 12
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192 Gene Cloning λ Clone Scaffold
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194 Gene Cloning Genomic DNA (a) Cl
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196 Gene Cloning therefore sequence
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198 Gene Cloning Tag sequence CGTGT
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200 Gene Cloning Genomic DNA (a) Ad
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204 Gene Cloning A7.3. (a) The dide
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Bioinformatics 209 Table 8.1 The ge
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Bioinformatics 243 A8.1. The triple
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Bioinformatics 245 Q8.11. Which gro
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250 Gene Cloning What sorts of stud
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252 Gene Cloning 9.2 Requirements f
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254 Gene Cloning (a) The lac promot
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256 Gene Cloning lac: CCAGGCTTTACAC
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258 Gene Cloning phage (such as T7)
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260 Gene Cloning 9.4 Some Problems
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262 Gene Cloning Figure 9.6 Inclusi
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264 Gene Cloning Box 9.1 Translatio
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266 Gene Cloning Box 9.2 Signal Seq
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268 Gene Cloning Ribosome Cytosol P
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270 Gene Cloning Other yeast specie
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272 Gene Cloning (turned on by an i
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274 Gene Cloning 9.6 A Final Word A
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10 Gene Cloning in the Functional A
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316 Gene Cloning genomic sequences
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318 Gene Cloning the first exon of
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320 Gene Cloning Box 11.1 End Label
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322 Gene Cloning mRNA 5' 3' 3' 5' S
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324 Gene Cloning mRNA 5' 3' 3' 5' G
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326 Gene Cloning cloning step. RACE
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328 Gene Cloning Prepare RNA from c
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330 Gene Cloning Gene A condition 1
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332 Gene Cloning (a) (b) (c) Immobi
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334 Gene Cloning tat and one in whi
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336 Gene Cloning Box 11.3 Examples
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338 Gene Cloning (a) MCS for promot
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340 Gene Cloning important regulato
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342 Gene Cloning (a) F DP DPA DNA-t
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344 Gene Cloning DNA alone DNA-prot
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346 Gene Cloning [FNR] GA - + -90 -
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348 Gene Cloning (a) 1 2 3 4 5 Prom
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350 Gene Cloning bind the transcrip
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352 Gene Cloning Target sequence Ye
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354 Gene Cloning have been develope
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356 Gene Cloning detected by autora
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358 Gene Cloning vide a complete li
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360 Gene Cloning MS 503 750 1400 16
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362 Gene Cloning Q11.3. Why is it e
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364 Gene Cloning (transcriptomics o
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366 Gene Cloning Figure 12.1 Transg
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368 Gene Cloning Herbicide Non-tran
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370 Gene Cloning that has been used
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372 Gene Cloning tumors with high f
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374 Gene Cloning Transgene encoding
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376 Gene Cloning placed on the mark
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378 Gene Cloning Gene A: a gene whi
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380 Gene Cloning Isolate the gene o
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382 Gene Cloning Box 12.1 Recombina
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384 Gene Cloning damage to the DNA
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386 Gene Cloning Box 12.2 The Produ
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388 Gene Cloning as to whether the
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390 Gene Cloning transformed. The i
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392 Gene Cloning 1. Gene of interes
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394 Gene Cloning Showing that the D
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396 Gene Cloning complete plants by
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398 Gene Cloning 12.5 Knockout Mice
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400 Gene Cloning introduced back in
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402 Gene Cloning Chromosome Constru
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404 Gene Cloning cells, and one has
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