Nonmammalian Genomic Analysis: A Practical GuideBruce Birren, Eric Lai Offering detailed protocols for those needing to construct a variety of maps and isolate genes, this unique book is intended to popularize the new techniques of genome analysis derived from the Human Genome Project. The power of these new methods is often most striking when applied to problems outside of human genetics, particularly the nonmammalian systems on which many researchers focus. Many of these organisms are economically important and biologically rich. Nonmammalian Genomic Analysis: A Practical Guide covers the "how to" aspects of preparation, handling, cloning, and analysis of large DNA and the creation of chromosome and genome maps. This lab manual facilitates the transfer of these technologies to small "low tech" environments and allows them to be used by those with no background in genome mapping or large-fragment cloning. Like having a local expert, this collection provides procedures for anyone, anywhere, and allows the replication of others' success.
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Page 63
... sterile water. Store at -20°C. (11) ES: 0.5 M EDTA, pH 9–9.5, 1% sarkosyl. Prepare as described for ESP. (12) Equilibration buffer: 10 mM Tris, 100 mM NaCl, 10 mM EDTA, adjust to pH 7.5 with 1 NHCl. Ill. Procedures A. Isolation of High ...
... sterile water. Store at -20°C. (11) ES: 0.5 M EDTA, pH 9–9.5, 1% sarkosyl. Prepare as described for ESP. (12) Equilibration buffer: 10 mM Tris, 100 mM NaCl, 10 mM EDTA, adjust to pH 7.5 with 1 NHCl. Ill. Procedures A. Isolation of High ...
Page 65
... sterile equipment. We use freshly bent pasteur pipettes for this purpose. Microbeads can be pipetted with cut-off pipet tips (3–5 mm tip diameter). Transfer blocks for restriction enzyme digestion into a 50-ml conical centrifuge tube ...
... sterile equipment. We use freshly bent pasteur pipettes for this purpose. Microbeads can be pipetted with cut-off pipet tips (3–5 mm tip diameter). Transfer blocks for restriction enzyme digestion into a 50-ml conical centrifuge tube ...
Page 66
... sterilized dialysis bag and dialyze the DNA at 4°C for 16 hr against 1–2 liters of TE 10/1 with one change of the buffer. (7) After dialysis the DNA can be stored at 4°C. DNA isolated in that manner is suitable for use in standard ...
... sterilized dialysis bag and dialyze the DNA at 4°C for 16 hr against 1–2 liters of TE 10/1 with one change of the buffer. (7) After dialysis the DNA can be stored at 4°C. DNA isolated in that manner is suitable for use in standard ...
Page 67
... sterile conditions. In our experience, two points are critical with in vitro-grown plants. One is that such plants usually contain large amounts of starch (see below). The other point is that leaves from such plants are very tender (and ...
... sterile conditions. In our experience, two points are critical with in vitro-grown plants. One is that such plants usually contain large amounts of starch (see below). The other point is that leaves from such plants are very tender (and ...
Page 94
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Contents
1 | |
25 | |
61 | |
Chapter 4 Generating and Using DNA Markers in Plants | 75 |
Chapter 5 Genome Mapping of Protozoan Parasites by Linking Clones | 135 |
Chapter 6 Macrorestriction Mapping and Analysis of Bacterial Genomes | 165 |
Chapter 7 Cosmid Cloning with Small Genomes | 197 |
Chapter 8 Construction of PI Artificial Chromosome PAC Libraries from Lower Vertebrates | 223 |
Chapter 9 The Selection of ChromosomeSpecific DNA Clones from African Trypanosome Genomic Libraries | 257 |
Chapter 10 Analysis of the Dictyostelium discoideum Genome | 293 |
Chapter 11 Integrated Genome Mapping by Hybridization Techniques | 319 |
Index | 347 |
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Common terms and phrases
Acad AFLP agarose gel aliquots amplification bacterial bacteriophage bands blot buffer cells centrifuge cerevisiae chDNAs chromosomes coli colonies containing contigs cosmid Dictyostelium DNA fragments DNA molecules DNA sequence EDTA electrophoretic karyotype ethidium bromide filters Final concentration gel electrophoresis gene Genet genomic DNA hybridization identified Incubate insert isolation karyotype lane large DNA ligation linear linking clones markers method mg/ml molecular NaCl Natl Novozym Nucleic Acids Res nucleotide oligonucleotide PAC cloning partial digestion PFGE physical mapping plasmid plates plugs polymerase polymorphisms prepare primers probe procedure proteinase K protocol protoplasts pulse pulsed-field gel RAPD reaction repeat restriction digestion restriction endonuclease restriction enzyme restriction fragments restriction mapping resuspend RFLP room temperature samples selected separation Sfil solution Southern blot specific spheroplasts sterile strains switch techniques tion Tris-HCl trypanosome tube V/cm vector vector DNA yeast yeast artificial chromosomes