Analysis Of Pcr Products

There are numerous methods for analysis of PCR products (Figs. 2 and 3). The method of choice for analysis of PCR products will depend on the type of information that is desired. Typical analysis of PCR products will involve electrophoretic separation of amplicons and visualization with ethidium bromide or other DNA dye. In most cases, amplification products can be analyzed using standard agarose gel electrophoresis. Agarose gel electrophoresis effectively separates DNA products over a wide range of sizes (100 bp to >25 kbp). PCR products from 200 to 2000 bp can be separated quickly on a 1.6% agarose gel. When greater resolution or separation power is required, such as in the analysis of very small PCR products (<100 bp), polyacrylamide gel electrophoresis is the method of choice. In both cases, ethidium bromide can be added to the gel before casting or the gel can be stained with ethidium bromide following electrophoresis. DNA products are easily visualized by ultraviolet illumination after ethidium bromide staining. Another method often used to quantify products is the incorporation of radioactive, fluorescent, or biotinylated markers. PCR products can be labeled by incorporating labeled nucleotides or

Fig. 3. Analysis and verification of PCR products. Amplicons produced using standard PCR can be analyzed and verified using various methods, some of which involve direct visualization of the PCR product (typically based on analysis of amplicon size). Other methods characterize the nature of the PCR product (based on sequence or sequence characteristics) or employ the PCR product in hybridization techniques.

Fig. 3. Analysis and verification of PCR products. Amplicons produced using standard PCR can be analyzed and verified using various methods, some of which involve direct visualization of the PCR product (typically based on analysis of amplicon size). Other methods characterize the nature of the PCR product (based on sequence or sequence characteristics) or employ the PCR product in hybridization techniques.

through the use of labeled oligonucleotide primers. Labeled PCR products are separated by electrophoresis on either agarose or polyacrylamide gels and visualized using appropriate techniques (such as autoradiography for radioactively labeled products).

In some cases, the desired information resulting from a PCR analysis can be obtained through a simple analytical gel separation, whereas in other instances, additional information is required. PCR products can be cloned and used for sequence analysis, construction of molecular probes, mutation analysis, in vitro mutagenesis, studies of gene expression, and many other applications. PCR fragments can be introduced into suitable vectors via several methods (15), where they can be expanded and/or manipulated. A cloning strategy should be planned prior to the PCR reaction because modifications of the product, such as the insertion of restriction sites, are sometimes necessary to allow insertion of the product into the intended vector. Care should also be taken to verify the size and purity of the PCR product prior to cloning. With careful planning and the inclusion of appropriate controls, the conventional techniques of recombinant DNA technology can be replaced almost entirely with PCR-based methods.

100 Pregnancy Tips

100 Pregnancy Tips

Prior to planning pregnancy, you should learn more about the things involved in getting pregnant. It involves carrying a baby inside you for nine months, caring for a child for a number of years, and many more. Consider these things, so that you can properly assess if you are ready for pregnancy. Get all these very important tips about pregnancy that you need to know.

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