What do you think?
Rate this book
The Polymerase Chain Reaction
James D. Watson When, in late March of 1953, Francis Crick and I came to write the first Nature paper describing the double helical structure of the DNA molecule, Francis had wanted to include a lengthy discussion of the genetic implications of a molecule whose struc ture we had divined from a minimum of experimental data and on theoretical argu ments based on physical principles. But I felt that this might be tempting fate, given that we had not yet seen the detailed evidence from King's College. Nevertheless, we reached a compromise and decided to include a sentence that pointed to the biological significance of the molecule's key feature-the complementary pairing of the bases. "It has not escaped our notice," Francis wrote, "that the specific pairing that we have postulated immediately suggests a possible copying mechanism for the genetic material." By May, when we were writing the second Nature paper, I was more confident that the proposed structure was at the very least substantially correct, so that this second paper contains a discussion of molecular self-duplication using templates or molds. We pointed out that, as a consequence of base pairing, a DNA molecule has two chains that are complementary to each other. Each chain could then act ". . . as a template for the formation on itself of a new companion chain, so that eventually we shall have two pairs of chains, where we only had one before" and, moreover, " ...
550 pages, Hardcover
First published September 30, 1989
59 people want to read
Ratings & Reviews
Friends & Following
Create a free account to discover what your friends think of this book!
Community Reviews
Displaying 1 of 1 review
Read
December 29, 2022Mullis tells a good story as he recounts his invention of PCR in 198
So by 1986, we could use in vitro systems to synthesize DNA with high efficiency, and because both DNA sequencing and oligonucleotide synthesis had become routine
December 1988, demonstrated that the polymerase chain reaction now ranked with cloning and DNA sequencing as an indispensable tool in the molecular biologist's armamentarium.
How slow progress would be without direct sequencing, mapping using micro satellite repeats, and sequence tagged sites.
The first paper on PCR dealt with the diagnosis of sickle cell anemia
Penrose's chapter on mathematical truth with its implication that certain things mayor may not be invented, but others were there already, and given time, would be discovered.
There are too many papers out there, and like technical papers tend to be when there are a lot of them all in one stack and they aren't yours, become so tedious that no one would live through reading them all anyhow!
There are two very distinct things that the polymerase chain reaction does. It generates a particular discrete DNA molecule that was not present to begin with, and it amplifies DNA molecules selectively.
Virtually any DNA sequence can also be engineered by "copying and pasting" with PCR as a replacement for conventional recombinant DNA technology where DNA is manipulated by "cutting and pasting" using restriction endonucleases and ligase
mutated and correct sequences are amplified at the same efficiency
The probability of producing fragments without error (p) in one cycle of amplification is given by the probability of no-hit in a Poisson distribution.
electrophoresis at 4 to lOoC in gel without glycerol usually results in a good separation. Addition of 5-10% glycerol to the gel and electrophoresis at 20-25°C will also give satisfactory results. It should be emphasized, however, that some mutations can be detected only under specific conditions (e.g., at 25°C without glycerol), although such mutations seem rare.
ethidium bromidestained gel
the presence of a few mutations is relatively unimportant
PCR conditions that reliably produce desired products result in a relatively high mutation rate (-1 % after 30 rounds).
PCR conditions that result in a minimum of mutations require the use of nucleotides (dNTPs) at low concentrations (Eckert and Kunkel, 1990). Using the conditions recommended for Taq polymerase by Perkin-ElmerCetus (0.2 mM) results in an error rate of -0.05% after 30 rounds of amplification.
rapid amplification of cDNA ends (RACE)
So by 1986, we could use in vitro systems to synthesize DNA with high efficiency, and because both DNA sequencing and oligonucleotide synthesis had become routine
December 1988, demonstrated that the polymerase chain reaction now ranked with cloning and DNA sequencing as an indispensable tool in the molecular biologist's armamentarium.
How slow progress would be without direct sequencing, mapping using micro satellite repeats, and sequence tagged sites.
The first paper on PCR dealt with the diagnosis of sickle cell anemia
Penrose's chapter on mathematical truth with its implication that certain things mayor may not be invented, but others were there already, and given time, would be discovered.
There are too many papers out there, and like technical papers tend to be when there are a lot of them all in one stack and they aren't yours, become so tedious that no one would live through reading them all anyhow!
There are two very distinct things that the polymerase chain reaction does. It generates a particular discrete DNA molecule that was not present to begin with, and it amplifies DNA molecules selectively.
Virtually any DNA sequence can also be engineered by "copying and pasting" with PCR as a replacement for conventional recombinant DNA technology where DNA is manipulated by "cutting and pasting" using restriction endonucleases and ligase
mutated and correct sequences are amplified at the same efficiency
The probability of producing fragments without error (p) in one cycle of amplification is given by the probability of no-hit in a Poisson distribution.
electrophoresis at 4 to lOoC in gel without glycerol usually results in a good separation. Addition of 5-10% glycerol to the gel and electrophoresis at 20-25°C will also give satisfactory results. It should be emphasized, however, that some mutations can be detected only under specific conditions (e.g., at 25°C without glycerol), although such mutations seem rare.
ethidium bromidestained gel
the presence of a few mutations is relatively unimportant
PCR conditions that reliably produce desired products result in a relatively high mutation rate (-1 % after 30 rounds).
PCR conditions that result in a minimum of mutations require the use of nucleotides (dNTPs) at low concentrations (Eckert and Kunkel, 1990). Using the conditions recommended for Taq polymerase by Perkin-ElmerCetus (0.2 mM) results in an error rate of -0.05% after 30 rounds of amplification.
rapid amplification of cDNA ends (RACE)
Displaying 1 of 1 review