The Polymerase Chain Reaction PCR

After the complementary strands are separated from each other, the primers attach themselves to the ends of the strands one primer becomes attached to one template strand, the other, to the complementary strand. Next, the enzyme causes bases to be added at the end of the primers, extending the formation of the complementary strands initiated by the 

At the end of the first cycle, which takes only minutes or an even shorter time, the number of DNA fragment is doubled. The cycle can be repeated indefinitely after 30 cycles, for example, which are completed in only a few hours, there will be about a billion copies of the original DNA fragment, sufficient for studying its nature and characteristics (see Fig. 76). 

Collecting samples of ancient nucleic acids is a delicate operation that requires what are basically surgical procedures. It is advantageous, whenever possible, that the samples be collected at excavation sites and precautions taken to ensure that they do not become contaminated with other, particularly more recent, nucleic acids. At high temperatures and humidity, nucleic acids decay quickly. Well-preserved ancient nucleic acids can, therefore, be expecfed in sifes where low femperafures and a dry environment prevail, as, for example, in cold, deserf areas of fhe world. Once collected, the samples need to be isolated from any ofher remaining maferials until they can be amplified by PCR and fheir chemical composition and sfructure can fhen be sfudied. 

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FIGURE 28.14 The polymerase chain reaction (PCR). Three cycles are shown the target region appears after the third cycle. Additional cycles lead to amplification of the target region. 

Sequencing of DNA is carried out by the Sanger dideoxy method, and small DNA segments can be synthesized in the laboratory by automated instruments. Small amounts of DNA can be amplified by factors of 106 using the polymerase chain reaction (PCR). 

While many diseases have long been known to result from alterations in an individual s DNA, tools for the detection of genetic mutations have only recently become widely available. These techniques rely upon the catalytic efficiency and specificity of enzyme catalysts. For example, the polymerase chain reaction (PCR) relies upon the ability of enzymes to serve as catalytic amplifiers to analyze the DNA present in biologic and forensic samples. In the PCR technique, a thermostable DNA polymerase, directed by appropriate oligonucleotide primers, produces thousands of copies of a sample of DNA that was present initially at levels too low for direct detection. 

There have been a number of attempts to achieve this objective, but so far the challenge has not been fully met. This Chapter will examine some of the conventional approaches and then go on to consider how recent developments in the use of the Polymerase Chain Reaction (PCR) with DNA for the identification of species and individual organisms by DNA analysis, sometimes known as DNA Fingerprinting", have identified a yet unrealized need for a new dimension of certified reference materials. 

Kasai K, Nakamura Y and White R 1990) Amplification of a variable number of tandem repeats (VNTR) locus (pMCTii8) by the polymerase chain reaction (PCR) and its application to forensic science. ] For Sci 35 1196-1200. 

Such methods also make it possible to learn about the state of health of the dead before death, the diseases from which they suffered, their age at the time of death, the method used for their mummification, and even the cultural environment in which they lived and were mummified (Cockbum et al. 1998 Harris and Wente 1980). The conception and development of the polymerase chain reaction (PCR) at the end of the twentieth century made it also possible to study the genetic characteristics of the mummies and of the populations to which they belonged (see Textbox 65). 

The same concept of volumetric in situ heating by microwaves was also exploited by Larhed and coworkers in the context of scaling-up a biochemical process such as the polymerase chain reaction (PCR) [25], In PCR technology, strict control of temperature in the heating cycles is essential in order not to deactivate the enzymes involved. With classical heating of a milliliter-scale sample, the time required for heat transfer through the wall of the reaction tube and to obtain an even temperature in the whole sample is still substantial. In practice, the slow distribution of heat... 

One of the in vitro (in the test tube) processes used to clone DNA is called the polymerase chain reaction (PCR). A vial in which PCR is to be carried out contains all the necessary components for DNA duplication the piece of DNA to be cloned large quantities of the four nucleotides, A, T, C, G large quantities of a primer sequence, a short sequence of about 20 nucleotides synthesized by the primase enzyme and DNA polymerase.To conduct the process, the vial is hrst heated to 90-95°C for 30 seconds to separate the two DNA chains in... 

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