Crick, Brenner et al. experiment
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The Crick, Brenner, Barnett, Watts-Tobin experiment of 1961 was a scientific experiment performed in 1961 by Francis Crick, Sydney Brenner, Leslie Barnett and R.J. Watts-Tobin. They demonstrated that three bases of DNA code for one amino acid in the genetic code. The experiment elucidated the nature of gene expression and frame-shift mutations.
In the experiment, proflavin-induced mutations of the T4 bacteriophage gene, rIIB, were isolated. Proflavin causes mutations by inserting itself between DNA bases, typically resulting in insertion or deletion of a single base pair.
The mutants produced by Crick and Brenner could not produce functional rIIB protein because the insertion or deletion of a single nucleotide caused a frameshift mutation. Mutants with two or four nucleotides inserted or deleted were also nonfunctional. However, the mutant strains could be made functional again by using proflavin to insert or delete a total of three nucleotides. This proved that the genetic code uses a codon of three DNA bases that corresponds to an amino acid.
Sydney Brenner, in his autobiographical book (with other contributors), describes the origin of this experiment on pages 91 to 96. He wrote: “It was possible to get a mutation in one place which would alter a protein, and then you could get another mutation elsewhere [in the same gene] which corrected the protein. It’s called an internal suppressor.” “Francis [Crick] started to play around in the lab with [rII] mutants. He started with a few of the base analogue mutants and these did not give him much joy. But where he found innumerable suppressors was with the acridine mutants [these were addition/deletion mutants]… You could start with a mutant which was arbitrarily called ‘plus’ because it gained a base. Then all the suppressors of it would be ‘minuses’ because when you add a plus and a minus you came back to zero….If you put [recombined] three [plus] mutants together [or three minus mutants together] they came back to normal! And if you mixed four or five they were still mutant. We went as far as six to make it normal and after that it got a bit boring.” “This concept of a phase shift, or a ‘frameshift’ [in the genetic code of an rII gene] as we later called it, was so foreign to people in genetics that we had endless problems trying to explain this work.”
The demonstration of the triplet nature of the genetic code, although carried out with bacteriophage, proved to be universally applicable to all forms of life. The experiment did not depend on knowing the sequence of DNA base pairs within the rIIB gene nor the amino acid sequence of its protein product, since this type of information was unobtainable in 1961. Rather the demonstration depended on the innovative use of frameshift (addition/deletion) mutants. Once it was established that the genetic code was a triplet code, and since it was known that there are only 4 types of base in DNA (A, T, G and C), it could be deduced that there are 64 possible triplets (codons) (4 x 4 x 4 = 64). With this understanding, it soon became possible for other investigators to assign each of 61 codons to a specific amino acid and three codons to polypeptide chain termination signals. These assignments, once established also proved to be nearly universal.
Crick FH, Barnett L, Brenner S, Watts-Tobin RJ (December 1961). "General nature of the genetic code for proteins". Nature 192 (4809): 1227–32. Bibcode:1961Natur.192.1227C. doi:10.1038/1921227a0. PMID 13882203.
1. Sydney Brenner (Author), Lewis Wolpert (Contributor), Errorl C. Friedberg (Contributor), Eleanor Lawrence (Contributor) 2001 My Life in Science: Sydney Brenner, A Life in Science 2001 Biomed Central Ltd (publisher) ISBN 0954027809 ISBN 978-0954027803
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