Polymerase chain reaction extraction

Fig. 19.1 Differential displays comparing RNAs from saline (S)-, imipramine (I)- or fluoxetine (F)-treated rats. Total RNA was extracted from hypothalami of animals treated with the different drugs for two months. Autoradiograms of amplified -[35S]-dATP-labeled PCR (polymerase chain reaction) products after electrophoresis in 6% polyacrylamide gels are shown for two different primer combinations that identified one upregulated (arrowhead) and one downregulated (arrow) fragment in the groups treated with antidepressants (from [4] with permission).

Shibata D. Extraction of DNA from paraffin-embedded tissue for analysis by polymerase chain reaction new tricks from an old friend. Hum. Pathol. 1994 25 561-563. 

Sato Y, Sugie R, Tsuchiya B, et al. Comparison of the DNA extraction methods for polymerase chain reaction amplification from formalin-fixed and paraffin-embedded tissues. Diagn. Mol. Pathol. 2001 10 265-271. 

The RNA molecules, ribosomal RNA (rRNA) and messenger RNA (mRNA) play key roles in the protein synthesis. The amount of RNA in individual cells or in a community may, therefore, be taken as an indicator of protein synthesis and, thus, microbial activity. The number of active cells can be detected by fluorescent in situ hybridization (FISH) (Amann et al. 1995). By this method, individual cells carrying high concentrations of rRNA, situated on ribosomes, are quantified by fluorescence microscopy. The amount of rRNA in a community can also be detected by Reverse Transcriptase Polymerase Chain Reaction (RT-PCR), where rRNA extracted from soil is detected by creating a DNA copy and separating by gel electrophoresis (Duineveld et al. 2001). 

Figure 2 Cytokine gene expression in immunopotentiating reconstituted influenza virosomes (IRIV) stimulated peripheral blood mononuclear cells (PBMC). PBMC were cultured in the presence or absence of IRIV. On days 1 and 2, culture cells were harvested and total cellular RNA was extracted and reverse transcribed. The cDNAs thus obtained were tested in real time polymerase chain reaction assays in the presence of primers specific for the indicated cytokine genes. Source From Refs. 6 and 9.

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... 

Venous blood (8 mL) was collected into 15-mL tubes containing 50 mmol/L disodium EDTA, and genomic DNA was extracted by the standard (phenol/chloroform) method. The genotype frequencies for GSTPl were determined by polymerase chain reaction (PCR)/RFLP-based methods using DNA extracted from peripheral lymphocytes. 

Suggest how the researcher might use the polymerase chain reaction (PCR) to detect the presence of the circular form of the deleted DNA in an extract of the protist. 

Erb, R. W. Wagner-Dobler.I. (1993). Detection of polychlorinated biphenyl degradation genes in polluted sediments by direct DNA extraction and polymerase chain reaction. Applied and Environmental Microbiology, 59, 4065-73. 

Forensic Science Introduction DNA Extraction Methods in Forensic Analysis Polymerase Chain Reaction in the Forensic Analysis of DNA... 

There are several different methods based on 16S rRNA sequences that can be used to study bacterial community structure. The first approach involves extracting bulk nucleic acids from samples and then using this material in subsequent molecular analyses. A large number of these methods involve the polymerase chain reaction (PCR). Using PCR, target gene sequences... 

Polymerase chain reaction (PCR) purification kit, gel extraction kit, DNA mini-prep kit. 

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