Restriction Enzymes
Table 9
Restriction Enzymes Recognizing Palindromic
Penta-, Hepta-, and Octanucleotide Sequences177Recognition sequence
comprising 5 bp
Recognition sequence
comprising 7 bpRecognition sequence
comprising 8 bpAflI ÷ G/GWCC a PpuMI RG/GWCCY b
AhaI ÷ CC/WGG RsrlI CG/GWCCG
BcnI ÷ CC/SGG
EcoRII /CCWGG
NotI GC/GGCCGCaListed in the table are commercially available restriction enzymes. For sequences that
are recognized by different isoschizomers (see Table 5), a + marks the name of the enzyme
sold by most manufacturers.
bR = A orG, S = G or C, W = A orT, Y = T or C.
cleavage sites is only possible if each restriction fragment for a given
enzyme is cut by one of the other enzymes. This information leads to
the linkage of neighboring sequences. Therefore, a series of single and
double digests with all chosen enzymes and enzyme combinations
followed by the determination of the length of the fragments produced
has to be carried out to construct a restriction map. Partial digests of
end-labeled DNA produced by one enzyme can be employed for the
alignment of individual sites. Alternatively, two-dimensional gel elec-
trophoresis can be used for the alignment of restriction fragments: the
cleavage products of the first digest are analyzed in the first dimen-
sion; separated fragments are then digested with another enzyme (or
digested to completion with the same enzyme when a partial digest
was performed) and analyzed in the second dimension. The second
cleavage reaction can easily be performed by diffusing the appropriate
buffer and the enzyme into the gel, and after the reaction is completed
facturers. In addition, in each box enzymes that recognize degenerate sequences are listed.
The table allows one to identify a restriction enzyme for any given palindromic tetra- or
hexanucleotide recognition sequence and to select restriction enzymes that produce a desired
5' or 3' projecting single-strand region after cleavage. For example, if one needs DNA to be
cleaved within the sequence AGATCT between A and G, one should look for
A/ .... T in the column on the left and for GATC on the row on top. This defines BgllI as the
restriciton enzyme needed. An inspection of the column designated by GATC shows that
BgllI produces a 5' projecting single-stranded region after cleavage of the DNA, which is
compatible with products of an MboI, BamHI, or BclI as well as some of the products of an
XholI cleavage, because XholI has a degenerate recognition sequence.