Recombinant DNA Method-Mutagenesis By: Jessica Green
Mutagenesis • Mutagenesis (the creation or formation of a mutation) can be used as a powerful genetic tool. By inducing mutations in specific ways and then observing the phenotype of the organism the function of genes and even individual nucleotides can be determined.
Some types of Mutagenesis • Directed Mutagenesis • Site-directed/Site-specific Mutagenesis • Mismatched Mutagenesis
Directed Mutagenesis • a largely discredited hypothesis proposing that organisms can respond to environmental stresses through directing mutations to certain genes or areas of the genome.
Site-directed Mutagenesis • Where a specific site in a cloned DNA needs to be altered in a precise, pre-determined way • Can be designed to create specific nucleotide substitutions, deletions, and so on
Two different forms of Site-specific Mutagenesis • 5’ add-on mutagenesis - Diagram on page 148 of the text - a new sequence or chemical group is added to the 5’ end of a PCR product • Mismatched primer mutagenesis - primer is designed to be only partially complementary
Mismatched Mutagenesis • Diagram pg. 147 of text; Figure 5.19 • Can create a desired point mutation at a unique predetermined site within a cloned DNA molecule • At the intended mutation site it bears a base that is complementary
References • http://humogef.ifj.edu.pl/zaklad/laborator.html (pictures) • Textbook • http://www3.interscience.wiley.com/cgi-bin/jhome/10009058?CRETRY=1&SRETRY=0