I1. Look up and (a) write out the restriction enzyme recognition sites for the e
ID: 271967 • Letter: I
Question
I1. Look up and (a) write out the restriction enzyme recognition sites for the enzymes Xbal and Spel. (b) Explain and diagram why the ends generated by these two enzymes are “compatible". (c) Ex pain why a mixed Xbal -Spel site can't be recut by either enzyme. (d) Do you think there is an enzyme other than Xbal and Spel that could cleave this mixed site? Why? (e) Consult this chart to see if you are correct: (https://www.neb.com/tools-and-resources/selection-charts/compatible-cohesive-ends-and- generation-of-new-restriction-sites). Explain the answer. (f Use the same chart to investigate whether there are any pairs of sites generated by compatible enzymes that can be cleaved by one of those enzymes. (g) Why is the inability of Xbal and Spel to recut a mixed site important to assembly standards in Synthetic Biology?Explanation / Answer
Xba 1 RE site-
5' T CTAGA 3'
3' AGATC T 5'
Spe 1 RE site-
5' A CTAGT 3'
3' TGATC A 5'
if you look at the 4 nucleotides of each strand they both are same .so they will join and form new DNA
5' T CTAGT 3'
3' AGATC A 5'
here bold are Spe1 RE fragment and other are Xba 1 RE fragment.
now the resulting DNA fragment contains no recognition sequence of Xba 1 and Spe 1 so they both will not cut the ligated product.
the enzyme which recognizes the following underlined sequence will cut the ligated product
5' TCTAGT 3'
3' AGATCA 5'
Bfa 1 will cut 5' C TAG 3' site. because it is 4 bp palindromic site.
to making a recombinant DNA by adding DNA digested by these two enzymes it is necessary that they will not cut the ligated DNA product otherwise the recombinant DNA will not be formed.