Methods in Enzymology Vol.155 Recombinant DNA, Part F

This chapter focuses on the methodology that is used to characterize recognition sequences and on the application of HgiCI (BanI) fragment ends in mismatch cloning rather than on enzyme purification procedures. The gliding bacterium, Herpetosiphon giganteus became one of the most intensively screened groups of organisms in the search for new restriction enzymes. Among the 10 strains tested, 17 enzymes could be found with seven different but related recognition sequences. This led to a hypothesis regarding the evolutionary relationship among these enzymes and that it could be a basis for a better understanding of the biochemical mechanism of restriction enzyme-deoxyribonucleic acid (DNA) target interaction. Among these enzymes, HgiCI is highly different from all other previously described endonucleases, because it produces 5'-hexanucleotide protruding ends. It is mentioned that BanI also produces 5'-hexanucleotide protruding deoxyribonucleic acid (DNA) fragments. Mixed ligase reactions are a quick and easy method to characterize an endonucleolytic cleavage reaction if a suspected isoschizomer is available. The chapter mentions the application of this technique for the AvaII/HgiCII cleavage site and used gel electrophoretic separation to identify the multiple ligation products in an agarose gel system.

• Series: Methods in Enzymology (Book 155)
• Hardcover: 628 pages
• Publisher: Academic Press;
• Language: English
• ISBN-10: 0121822664
• ISBN-13: 978-0121820565
• ASIN: 0121820564
• Product Dimensions: 1.2 x 6 x 9.2 inches

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