Restriction Map
Map the cut sites of 109 common restriction enzymes on a linear or circular DNA sequence, find the single cutters and the enzymes that don't cut, then pick the enzymes for a digest and see its fragments, their sizes and how they would run on a gel.
Sequence tool
Map the cut sites
Bare letters, a FASTA record or a GenBank record. Numbers and spaces from a listing are ignored.
Paste a DNA sequence or a plasmid to map where the enzymes cut.
Worked example
pUC19 (2,686 bp, circular) cut with EcoRI and HindIII
- 01EcoRI, G^AATTC, at 396: cut after 396
- 02HindIII, A^AGCTT, at 447: cut after 447
- 03two cuts on a circle: 447 − 396 = 51 bp, and 2,686 − 51 = 2,635 bp
Two fragments, 51 bp and 2,635 bp: the polylinker drops out, and the vector backbone is left with an EcoRI end and a HindIII end, ready for an insert cut the same way.
How it works
The reasoning behind it
A restriction enzyme recognises a short sequence, usually four to eight bases and usually a palindrome, and cuts both strands at a fixed place in or near it. The tool searches the sequence for each enzyme's site, on both strands for the enzymes whose sites aren't palindromes, and lists every cut.
The cuts you choose for a digest are sorted along the sequence, and the pieces between them are the fragments. A linear molecule with n cuts gives n + 1 fragments; a circle gives n, and with one cut it opens into a single full-length line.
The gel is drawn from the fragment sizes: a fragment's distance down the lane follows the logarithm of its length, as it does in agarose, beside a 1 kb ladder for comparison. Two fragments of nearly the same size land on the same band, which is why a digest sometimes shows fewer bands than fragments.
Questions
Frequently asked
How do I find an enzyme that cuts my plasmid once?
Map it with every enzyme and look at the single cutters list. For cloning, choose one that cuts in the polylinker and nowhere in the insert, and check the insert's map too.
Why does the map show a site my digest doesn't cut?
Most often methylation: DNA from a normal E. coli strain carries Dam and Dcm methyl groups that block some enzymes at some sites. Otherwise the enzyme may be old, the buffer wrong, or two sites so close that one blocks the other.
What is the difference between a 5′ overhang, a 3′ overhang and a blunt end?
It is which strand is cut first. EcoRI cuts the top strand before the bottom, so each end has a single-stranded 5′ tail (AATT). PstI cuts the bottom strand first and leaves a 3′ tail (TGCA). SmaI cuts both in the same place and leaves no tail. Ends ligate to ends with a matching tail, or blunt to blunt.
Which enzymes are included?
109 enzymes sold by the main suppliers: the common six- and eight-base cutters, the four-base cutters used for fingerprinting, and the type IIS enzymes used in Golden Gate assembly. Sites and cut positions are from REBASE and were checked against Biopython on pUC19.
Why do I get one fragment fewer than I expected?
Because the sequence is circular. A circle with two cuts gives two fragments, not three. If you pasted a linear sequence, switch the setting, and the end pieces appear.
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