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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From inside the book
Results 1-5 of 41
Page 12
... EDTA 25 9.3 g disodium EDTA-2 H2O H2O - Add to bring final volume to 1 liter The pH of this mixture will be 8.3. 2. l X TAE Amount needed to prepare 1 liter of a 50X Component Final concentration (mM) concentrated solution Tris 40 242 g ...
... EDTA 25 9.3 g disodium EDTA-2 H2O H2O - Add to bring final volume to 1 liter The pH of this mixture will be 8.3. 2. l X TAE Amount needed to prepare 1 liter of a 50X Component Final concentration (mM) concentrated solution Tris 40 242 g ...
Page 14
... EDTA (used to inhibit endogenous nucleases that could degrade the DNA during the incubations). This is usually the only treatment necessary for organisms that lack a cell wall. Digesting a cell wall usually requires an additional step ...
... EDTA (used to inhibit endogenous nucleases that could degrade the DNA during the incubations). This is usually the only treatment necessary for organisms that lack a cell wall. Digesting a cell wall usually requires an additional step ...
Page 15
... EDTA, pH 8.0, to wash the cells. Collect cells by spinning again for 5 min as before and decant supernatant. (4) Resuspend cells in 6 ml 50 mM EDTA, pH 8.0, and mix in 160 ml 10 mg/ml Zymolyzase 201. (5) Briefly warm cells to 37°C and ...
... EDTA, pH 8.0, to wash the cells. Collect cells by spinning again for 5 min as before and decant supernatant. (4) Resuspend cells in 6 ml 50 mM EDTA, pH 8.0, and mix in 160 ml 10 mg/ml Zymolyzase 201. (5) Briefly warm cells to 37°C and ...
Page 16
... EDTA. (5) Collect the cells by centrifuging at 4000g for 10 min. (6) Discard supernatant and resuspend cells in 0.5 ml of 200 mM NaCl, 10 mM Tris, pH 7.2, 100 mM EDTA by pipetting. Briefly warm to 37°C. (7) Add an equal volume of 1% low ...
... EDTA. (5) Collect the cells by centrifuging at 4000g for 10 min. (6) Discard supernatant and resuspend cells in 0.5 ml of 200 mM NaCl, 10 mM Tris, pH 7.2, 100 mM EDTA by pipetting. Briefly warm to 37°C. (7) Add an equal volume of 1% low ...
Page 17
... EDTA prior to use. C. Controlling and Determining DNA Concentration The final concentration of DNA in the agarose is determined by the initial concentration of cells in the liquid agarose. However, once the agarose has solidified and ...
... EDTA prior to use. C. Controlling and Determining DNA Concentration The final concentration of DNA in the agarose is determined by the initial concentration of cells in the liquid agarose. However, once the agarose has solidified and ...
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