Restriction analysis

Much of what we know about the regulation of information flow (gene expression) has been made possible by the ability to manipulate the structures of DNA, RNA, and proteins and see how this affects their function. The ability to manipulate DNA (recombinant-DNA methods) has generated a new language filled with strange-sounding acronyms that are easy to understand if you know what they mean but impossible to understand if you don t. Understand  

Restriction enzymes are sequence-specific endonucleases that cut double-stranded DNA at specific sites. 

Most useful restriction enzymes cut DNA at specific recognition sites, usually four to six nucleotides in length. There can be multiple restriction sites for a single endonuclease within a given piece of DNA, there can be only one (a unique restriction site), or there can be none. It all depends on the sequence of the specific piece of DNA in question. 

Cutting with restriction endonucleases is very useful for moving specific pieces of DNA around from place to place. It s also a useful way to name pieces of DNA. For example, a piece of DNA that is cut from a bigger piece of DNA is often named by size and given a surname that corresponds to the two restriction enzymes that did the cutting—the 0.3-kb EcoRI-BamHI fragment. Restriction enzymes themselves are named for the bacterial strains from which they were initially isolated. 

A restriction map shows the location of restriction sites in a given DNA sequence. 

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DNA restriction analysis of the corresponding cosmids revealed one common c oRJ fragment of 9.2 kbp. The restriction map of the DNA region covered by the overlapping clones is shown in Fig 1. 

Presented below are four increasingly stringent confirmatory techniques for PCR and a brief discussion of considerations, limitations and advantages of each. These four techniques are agarose gel electrophoresis, restriction analysis. Southern blotting and sequencing. 

Infrared spectroscopy, including FTIR Polyacrylamide gel electrophoresis (PAGE) Amplified ribosomal DNA restriction analysis (ARDRA)... 

Restriction analysis can be used to detect sickle cell disease prenatally, since the DNA of all cells, including amniotic cells, carries the mutant DNA. It is much more difficult to obtain fetal blood for the analysis of the mutant hemoglobin A P-chain. Furthermore, fetal blood is composed mostly of fetal hemoglobin, sinee-hemoglobin A is made later in development. 

Newton, L.A., Chilton, N.B., Beveridge, I., Hoste, H., Nansen, P. and Gasser, R.B. (1998a) Genetic markers for strongylid nematodes of livestock defined by PCR-based restriction analysis of spacer rDNA. Acta Tropica 69,1-15. 

Provided a sample of DNA can be obtained, a restriction analysis can be carried ont. A match between the restriction fragments from a sample of DNA left at the scene of a crime and that of a snspect is a valnable tool in forensic science. The usefulness of this techniqne is increased enormously by combining it with the polymerase chain reaction, since the amount of DNA extracted from a very small amount of tissue can be increased enormonsly, providing enough for a restriction analysis. Tissne samples as small as a single cell, a hair, a drop of sahva, a piece of dandruff or a smear of semen are snfflcient to prodnce enough DNA. It has produced a revolution in forensic science. However, caution must be applied to interpretation of the results for... 

Hooper, S. W., Dockendorff, T. C. Sayler, G.S. (1989). Characteristics and restriction analysis of the 4-chlorobiphenyl catabolic plasmid, pSS50. Applied and Environmental Microbiology, 55, 1286-8. 

Analyze an FITC smgle-stained tube, such as FITC-CD8, scatter gating to restrict analysis to lymphocytes... 

Restriction Analysis of human genomic DNA has revealed that there are many differences... 

Item Isozyme electrophoresis MCF DNA-DNA hybridization Restriction analysis RAPD Sequencing PCR Cloning... 

Many workers have previously devoted attention to the contribution of errors in measurements to the problem of building trees from distances, as summarized in the contribution by Marshall.25 By contrast, we have not been concerned with this relatively minor source of error. Instead, our concern has been with a bigger source of error, the calibration error, which reflects the uncertainty in the relationship of distance measured with an indirect method to that measured with a more direct method. This aspect has not been addressed by previous workers. To illustrate the magnitude that this problem can assume, we note that DNA hybridization led to an estimate of 3.3% sequence divergence between the mitochondrial DNAs of two flies (Drosophila yakuba and D. teissieri).26 Restriction analysis done on the whole mitochondrial genome, in contrast, led to an estimate of 0.22%.27 Sequencing of one-seventh of these fly mitochondrial DNAs produced an estimate of 0.3%,27 similar to the latter indirect estimate but in dramatic contrast to the estimate from hybridization. 

Many of the new pesticides are active at low application rates, resulting in the need for sophisticated instrumentation for the traditional residue analysis. Such instrumentation is lacking in developing countries thus restricting analysis to older pesticides. The analytical capability of laboratories in developing countries needs to be improved in order that both research and monitoring can carried out on the environmental and health risks associated with the newer pesticides in tropical regions. 

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