Reference no: EM133240836

The restriction enzyme Sau3A recognizes the sequence 5'-/GATC-3' and cleaves on the 5' side of the G as indicated by the slash. Since the top and bottom strands of this restriction site are identical in the 5'→3' direction, only one strand of the site needs to be shown. The single-stranded ends produced by Sau3A cleavage are identical to those produced by BamHI, which recognizes the sequence 5'-G/GATCC-3' and cuts at the location of the slash. This means that an end cut by one enzyme can be ligated onto an end cut by the other enzyme by DNA Ligase. Given this information, answer the following questions:

A) Write out both strands of the recognition sequences for BamHI and Sau3A, show what they would look like after they are cut, and what the DNA would look like after you ligate a BamHI site to a Sau3A site. Make sure that it is easy to follow the individual pieces of DNA. You may want to use two different colors to keep them separate.

B) What fraction of BamHI sites can be cut by Sau3A? What fraction of Sau3A site can be cut by BamHI?

C) If two BamHI sites are ligated together, the resulting site can be cleaved again by BamHI. The same is true of two Sau3A ends. Suppose you ligate a fragment with a Sau3A end into a plasmid that has been cut by BamHI, can the hybrid site be cut with BamHI? Can it be cut with Sau3A?

D) Restriction enzyme recognition sites occur randomly throughout the genome. Calculate the probability of a Sau3A site (each nucleotide has 4 possibilities and in statistics the probability of multiple events is the probability of one multiplied by the probability of the other). Is that greater or less than the probability of a BamHI site? What is the probability of a BamHI site?