Genetic Manipulation: Techniques and Applications

394 Pages·1990·20.809 MB·English

THE SOCIETY FOR APPLIED BACTERIOLOGY
TECHNICAL SERIES NO 28
Genetic Manipulation
TECHNIQUES AND
APPLICATIONS
Edited ~)l
J. M. GRANGE
NationalHeart andLung11lStitute
Universi~)l ofLmuum
London
A. FOX &
N. L. MORGAN
South BankPolVlet/mit
London
OXFORD
BLACKWELL SCIENTIFIC PUBLICATIONS
LONDON EDINBURGH BOSTO"l
MELBOURNE PARIS BERLIN \IENl\A
Genetic Manipulation: Techniques and Applications Edited by J. M. Grange, A. Fox and N. L. Morgan
© 1991 the Society for Applied Bacteriology ISBN: 978-0-632-02926-6
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lransllliucd,indTl\ !(Jflllorb\al1\"means,electronic, (J-encticmanipulation:techniquesandapplications/
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withoutthepriorpermissionllfthecopyrighto.....vncr. p, ern,-(TheSocietyt<,rApplied
Bactcri{)l()g-~technicalserie..;no.2H)
brstpublished1991 "BaseduntheSoeietvofAppliedBacteriolog,
AutumnVleetingheldattheSouthBank
SctbySetritcTypesetters,HongKong Polytechnic,London~un IROctober1989"
PrintedandboundinGreatBritain Pref.
atthc'\ldcnPress,Oxj()rd Includeshihli()graphicalreferences.
andhoundatthe(~rcr:nStreet lncltldcsindex.
Bindery,(hfrm:..! ISBI'\0-632-02926-9
1. Geneticengineering-Congresses.
BritishLibrar- 2. Biotechnology---Congressl's. I. Grange,
Cataloguingil~Puhlicatlon1)<1ta JohnVI. II. Fox,.\,(\rnold),1927
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Contents
Contributors...... .......................................... Xl
Preface. .... ..... .......................................... xv
Extraction,Purificationand AssayofDNA .
KAREN E. WEBB AND R. WII.SON
IsolationofDNA, 1
IsolationofPlasmid D!'\A from Bacteria, 3
Purification ofD!'\A, 5
Isolating Genomic DNA from Bacterial Cells, 6
IsolatingMallllllalian DNA, 7
Assay ofDNA, 10
References, 13
RestrictionMappingofDNA ................................ 15
KAREN E. WEBB AND R. WII.SOK
Restriction Enzymes, 15
Approaches to ""lapping, 16
References, 23
The Construction and Use ofCloningVectors. ................ 25
P. MULLANY, B. WREK, M. WILKS, C.L. CLAYTON AND
SOAD TABAQCHALI
The Commercially Available Vectors, 25
The Construction ofVectors for Use with the Clostridia, 33
References, 35
Appendb( I: Addresses ofSuppliers ofCloning Vectors, 35
Appendb< II: Addresses ofSuppliers ofOther Products Useful for
Molecular Biology, 36
Adaptor Based eDNA Cloning in the Phage Vectors
AgtlO and Agtll 37
C.J. CARTER AND A.]. HAYLE
E:\:perimental Methods, 38
Results ofcDNA I-ibrary Analysis, 49
Discussion, 52
Acknowledgements, 53
References, 54
iii
IV CONTENTS
Plasmid Profiling and DNA/DNA Hybridization for
DistinguishingBetweenMesophilic AeromonasBacteria 55
S. ROGERS, M.F. MOFFATT, I.F. CONNERTON AND
R.W.A. PARK
DNA/DNA IIybridization, 55
Plasmid Profiling, 57
Methods, 58
Results, 62
Acknowledgements, 64
References, 65
Appendix: Reagents, 68
Preparation and ScreeningofBacterial Genomic Libraries 71
J.W. DALE
General Strategy, 71
Construction ofa Genomic Library by using f.ElVlBL4, 77
Screening ofa A.EMBL4 Library with Gene Probes, 84
Construction ofa ),gtll Libraryand Screeningwith Antibodies, 89
Acknowledgements, 95
References, 95
DNAProbes for Detection and IdentificationofBacteria 97
J.J.
McFADDEN AND A. KNIGHT
Principle ofNucleic Acid Hybridization, 97
Application ofDNA Hybridization, 99
The Polymerase Chain Reaction, 108
References, 109
Extractionand Purification ofEukaryoticmRNA ... .... . ...... 113
P.G. SANDERS, T. JACKSON, JANE NEWCOI\IBE,
SARAH BELL, G.E. SCOPES AND A. KNIGHT
Basic Precautions Prior to RNA Extraction, 114
RNA E:\traction Protocols, 115
Analysis ofPurified RNA, 122
Isolation ofPolyadenylated mRNA fi'om Total RNA, 123
Storage ofRNA Samples, 126
Acknowledgements, 126
References, 126
cDNA Cloning. ................................ ..... . . ..... 129
P.G. SANDERS AND A.J. EAST()N
How 'Viany Clones are Required?, 129
Cloning Strategies, 130
First-strand Synthesis, 134
Second-strand Synthesis, 137
Preparation ofthe cD:\'A for Cloning, 139
CONTENTS v
Methylation ofthe eDNA, 141
Linker Kinase Reaction, 141
TestofLigation Efficien(;)', 143
Ligation ofthe Phosphorylated Linkers to the eDNA, 143
EcoRI Digestion and Removal ofExcess Linkers from the cDNA, 143
Size Fractionation ofthe Linkered eDNA, 144
Ligating Fragments into pUC Vectors, 145
Transformation ofHost Bacteria, 146
Screeningfor Recombinant Phages and Plasmids, 147
References, 147
PrimerExtensionSequencingofRNAViruses. ................ 149
C.A. GIBSON, L.M. DUNSTER AND A.D.T. BARRETT
Precautions when Handling RNA, ISO
Nucleotide SequencingofInfluenza Virus Genes, lSI
Primer Ex1ension SequencingofInfluenza AViruses, 153
Primer Extension SequencingofFla\ivirus Genes, ISS
Comments, 161
References. 165
Appendix: Buffers and Reagents, 166
ForensicApplicationsofDNAProfiling....................... 169
B.H. PARKIN
Multilocus Probes, 170
Single Locus Probes, 171
Methods, 174
DNAProfiling in Forensic Casework, 175
References, 178
Application ofNucleic Acid Probes to the Identification of
BacterialEntericPathogens ................................. 181
B. \VREN, H. KU:ANTHOUS, C.L. CLAYTON,
P. MULLANY AND SOAD TABAQCHALI
Identification ofEnterotoxigenic Escherichia <viifrom Faecal Specimens, 182
Gene Probes for otherBacterial Enteric Pathogens, 184
Prospect~ for Nucleic Acid Probes in Clinical Microbiology, 185
Polymerase Chain Reaction Amplification ofDNA, 186
References, 188
Quantification ofCollagenmRNALevels inMammalianCells 191
J.S. CA:vtPA, ].E. BISHOP AND G.J. LAURENT
Preparation ofTotal RNA from Cells in Culture, 192
Quantification ofSpecific mRNA Levels, 195
References, 201
vi CONTENTS
Detection ofFoodbome Bacterial Pathogens by a
ColorimetricDNAHybridizationMethod. .................... 203
M. MOZOLA, D. HALBERT, S. CHAN, H-Y. lIsc,
A. JOllNSOI\, W. KING, S. WILSON, R.P. BETTS,
PAMELA BANKES AND J.G. BANKS
Colorimetric DNA IIybridization Assay System, 204
Conclusions, 213
References, 214
Use ofa DNAProbe for aViroidin PlantPathology. ........... 217
P.S. HARRIS AND C.M. JAMES
Materials and Methods, 218
Results, 221
Discussion, 223
Acknowledgements, 224
References, 224
Analysis ofRestriction Fragment Length Polymorphisms
in the StudyofBacteria ..................................... 227
N.A. SACNDERS
Epidemiolobrical Typing, 227
1Vlethods ofRFLP Analysis, 229
Examples ofResults, 240
Concluding Comments, 243
References, 243
The Use ofDNA Probes to Detect 'Selected Bacteria' After
TheirIntroductioninto aWasteWaterTreatmentPlant. ....... 245
GABY GAYER-HERKERT AND H.J. KCTZNER
;\1.aterials and iVlethods, 245
Results and Discussion, 248
Conclusions, 248
Acknowledgements, 251
References, 251
The Use ofGeneticManipulation to StudyBacterial Pathogens 253
R. WILSON AND KAREN E. WEBB
AGenetic Analysis ofPneumolysin (a ~irulence determinant ofStreptococcus
pneumoniae), 255
The Effect ofPseudomonas aemginosa on Respiratory Epithelium, 256
Methods, 260
References, 263
CONTENTS ~i
RNA DetectionMethods inBasic CardiacResearch. ........... 265
P.].R. BARTON, W.]. VALLINS, G.E. LYONS, N.]. BRAND
AND M. YACOUB
IsolationofRNA and mRNA Detection by Northern Blotting, 266
Probes for Hybridization Experiments, 267
In situ Hybridization, 271
References, 276
The Polymerase Chain Reaction and its Application to Basic
ResearchinMolecularBiology. .............................. 279
N.]. BRAND, W.]. VALLINS, M. YACOUB AND
P.].R. BARTON
The Polymerase Chain Reaction, 279
Methods, 281
Applications to Basic Research, 282
Conclusions and Perspectives, 289
References, 290
Appendix, 291
Problems and Troubleshooting, 291
The Effect ofNutrient Limitation on the Stability and
ExpressionofRecombinantPlasmids 295
A. WARl\ES AND].R. STEPHEl\SOl\
Structural Instability, 295
Seb'Tegational Instability, 296
Experimental Techniques used to Monitor Plasmid Stability, 299
Detection ofRecombinant Proteins, 302
Estimation ofthe Nutritional Requirements ofthe Host Organism, 306
Monitoring the Stability ofPlasmids Expressing Staphylococcal Protein A
in E. coli, 307
Summary, 310
Acknowledgements, 311
References, 311
Large-Scale Production ofProteins from Recombinant DNA in
Escherichiacoli ............................................ 315
R.F. SHERWOOD, R. PLAl\K, ]. BAKER AND T. ATKINSON
Genetic Design, 315
Fermentation, 322
Downstream Processing, 326
Quality Assurance, 332
Conclusion, 334
References, 334
viii CONTE:"lTS
Stabilityand CopyNumberofYeastCloningVectors. .......... 337
E.B. GINGOLD, M.J. KU:Il\lMAN AND VIRGINIA BUGEJA
Gene Cloningin Yeasts, 337
Plasmid Stability, 339
CopyNumber Determination, 343
References, 348
Designer Genes - Design and Synthesis Illustrated for the
Gene Coding for Human Macrophage Colony Stimulating
Factor " ... ... ............... 351
LINDA DAVIES, J.D. JOHNSON AND lA. DAVIES
Gene Design, 351
Materials and Methods, 353
Gene Synthesis, 355
Discussion, 358
References, 359
MonitoringSafetyinProcess Biotechnology. .................. 361
A.M. BENNETT, S.E. HILI., J.E. BENBOUGH AND
P. HArYIBLETON
MedlOds oflVlicrobial Air Monitoring, 362
Microbial Monitoringofa Bioprocessing Unit, 365
Results and Discussion, 366
Conclusions, 374
Acknowledgements, 375
References, 375
Patentingin Biotechnology. ................................. 377
D.L. WOOD
Fundamentals ofthe Patent System, 377
Biotechnological Inventions, 382
Obtaining a Patent, 386
Conclusions, 386
Index. ........ .................... ..... ............ ...... .. 388
List ofTitles in the Society for Applied Bacteriology Technical
Series. ....... . ............................................ 400
Contributors
T. ATKINSO.\l, Division of Biotechnology, Public Health Laboratory Semice,
CentrejOrAppliedMicrobiology andResearch, Porton Down, Salisbury, Wiltshire
SP40JG, UK
J. BAKER, Division ofBiotechnology, PublicHealth LaboratorySemice, CentrejOr
AppliedMicrobiologyandResearch, Porton Down, Salisbury, WiltshireSP4 OJG,
UK
PAMELA BANKES, Campden Food and Drink Research Association, Chipping
Campden, Gloucestershire GLSS 6LD, UK
J.G. BANKS, Campden FoodandDrinkResearchAssociation, ChippingCampden,
Gloucestershire GLSS 6LD, UK
A.D.T. BARRETT, DepartmentofiHicrobiology, Universi(y ofSurrey, GuildjOrd,
Surrey GU2 SXH, UK
P.J.R. BARTON, Department of Cardiothoracic Surgery, National Heart and
Lung Institute, Drroehouse Street, London SW3 6LY, UK
SARAH BELL,MolecularBiology Group, DepartmentofMicrobiology, Universi(yof
Surrey, Guildj()rd, Surre.y GU2 SXH, UK
J.E. BEMlOUGll, Division ofBiologics, PublicHealth LaboratorySemice, Centre
j(JrAppliedMicrobiology and Research, Porton Down, Salisbury, Wiltshire SP4
OJG, UK
A.1\1. BEN.\IETT, Division ofBiologics, PublicHealth Laboratory Service, Centre
jOrAppliedMicrobiology and Research, Porton Down, Salisbury, Wiltshire SP4
OJG, UK
R.P. BETTS, CampdenFoodandDrinkResearchAssociation, ChippingCampden,
Gloucestershire GL5S 6LD, UK
J.E. BISHOP, Biochemistry Unit, Nationailleart and LungInstitute, Dovehouse
Street, London SW3 6LY, UK
N.J. BRAND, Department ofCardiothoracic Surgery, National Heart and Lung
Institute, Dovehouse 5'treet, London SW3 6LY, UK
VIRGINIA I3UGEJA, Department of Biological Sciences, Hatfield Pozytechnic,
Hatfield, Hertjordshire ALl0 9A.B, UK
J.S. CA'VIPA, Biochemistryl Unit, National Heart and Lung Institute, Dovehouse
Street, London SW3 6L}~ UK
C.]. CARTER, Lift Sciences Division, Amersham InternationalPLC, W7zite Lion
Road, Amersham, Bucks HP7 9LL, UK
lX
x CONTRIBUTORS
S. CHAN, GENE-TRAK Systems, 31 New York Avenue, Framingham, Mas
sachusetts 01701, Uc~'A
C.L. CLAYTON, Department of Medical Microbiolof!J', St Bartholomew's
Hospital, London EC1A 7BE, UK
I.F. CONNERTOl\, Department ofMicrobiolof!J', Universi~y ofReading, London
Road, Reading, Berks RG1 SAQ UK
J .W. D ALE, MolecularBiolof!J' Group, Department ofMicrobiolof!J', Universi~y of
Surrey, GuildfOrd, Surrey GU2 SXH, UK
J.A. DAVIES, British Biotechnolof!J' Ltd, Watlington Road, Cowley, OxfOrd
OX4 SLY, UK
LINDA DAVIES, British Biotechnolof!J' Ltd, Watlington Road, Cowley, Oxjord
OX4 SLY, UK
L.M. DUNSTER, Department ofMicrobiolof!J', Universi~y ofSurrey, GuildfOrd,
Surrey GU2 5XH, UK
A.J. EASTON, Departmento.fBiologicalSciences, University o.fWarwick, Coventry
CV47AL, UK
G ABY GAYER- HERKERT, Institut jilr Microbiologie, Technische Hochschule
Darmstadt, Schnittpahnstmsse 10, 6100 Darmstadt, Germany
C.A. GIBSON, Department ofMicrobiolof!J', Universi~y ofSurrey, Guildford,
Surrey GU2 SXH, UK
E.B. GI'\IGOLD, Department ofBiotechnolof!J', South Bank Po~ytechnic, London
SEI0M, UK
D. HALBERT, GENE-TRAK c~)stems, 31 New York Avenue, Framingham,
Massachusetts 01701, USA
P. HAlvlBLETON, Division of Biologics, Public Health Laboratory Service,
CentrefOrAppliedMicrobiolof!J' andResearch, Porton Doum, Salisbury, Wiltshire
SP40JG, UK
P.S. HARRIS, Department ojAgriculture andFisheriesjor Scotland, Agricultural
Scientific Services, East Craigs, Edinburgh EH12 8NJ, UK
A.J. HAYLE, Life Sciences Division, Amersham International PLC, White Lion
Road, Amersham, Bucks HP7 9LL, UK
S.E. HILL, Division ofBiologics, Public Health Laboratory Service, Centre jor
AppliedMicrobiolof!J' andResearch, Porton Down, Salisbury, WiltshireSP4 OJG,
UK
II-Y. Hsu, GENE-TRAK c~ystems, 31 NelT! York Avenue, Framingham,
Massachusetts 01701, USA
T. JA.CKSON, A10lecular Biolof!J' Group, Department ofjt1icrobiolof!J', Universi~y
ofSurrey, GuildfOrd, Surre,y GU2 SXH, UK
C.M. JA.'VIES, Department ofAgriculture andFisheriesfor Scotland, Agricultural
Scientific Services, East Craigs, Edinburgh EH12 8NJ, UK
A. JOII.'iSO!\, GENE-TRAK Svstems, 31 NeuJ York Avenue, Framingham,
Massachusetts 01701, U5>1