RestrictionMapper Help
RestrictionMapper is a web site that finds restriction endonuclease cleavage sites in DNA sequences. It supports linear and circular DNA and provides several ways to sort and filter output. Also provided is a virtual digest function that simulates a simultaneous digest of your sequence with enzymes of your choice.
RestrictionMapper Directions
In the Conformation box, tell RestrictionMapper whether your DNA is linear or circular.
The Sort By lists let you sort your output 3 levels deep. "Frequency" means sort numerically by number of cuts. "Name" means sort alphabetically by name. "Site_length" means sort numerically by number of unique bases in the recognition sequence. "Overhang" means sort by overhang in this order: blunt, 5', 3'. If you choose 1: frequency, 2: site_length, 3: overhang, your table will first be sorted by number of cuts. Within each cut number (1, 2, etc.) the table will be sorted by site length. Each site length group will in turn be sorted by overhang.
In the Include section, choose what types of restriction enzymes you want in your output. The "NEB Only" option only returns enzymes sold by New England Biolabs.
The overhang check boxes let you filter the output by overhang. For example, to choose only blunt ended sites, uncheck the "5' overhang" and "3' overhang" boxes.
The Maximum Cut Number list lets you filter the output by the number of cut sites. For example, to find only enzymes that linearize your plasmid, select '1' (also don't forget to click 'circular'). To show only noncutters, select '0'.
The Minimum Site Length box lets you filter out enzymes with recognition sites below a certain length.
The Prototypes/All Isoschizomers radio buttons let you filter out isoschizomers.
The Select Enzymes by Name list lets you choose exactly which sites to search for. Control-click to select multiple enzymes. This list overrides all other selection criteria.
Your sequence is pasted into the Paste Sequence Here box.
Make sure that:
- Your sequence is single stranded. RestrictionMapper doesn't support, or
even recognize, double strands, so a double stranded sequence will give
you very strange results.
- There are no non-nucleotide letters in your sequence. Deal with any Ns
and remove the header before pasting. Numbers, spaces and blank lines are
no problem. RestrictionMapper removes these automatically.
The Sequence Name box is pretty self explanatory. The name is included in the output so that when you're looking at the printout 2 weeks later, you know what it is. The default name is "Untitled".
Now just click "Map Restriction Sites" and you're all set. RestrictionMapper will return the sequence name, the conformation of your DNA (linear or circular), your enzyme selection criteria, the list of non-cutters, and an enzyme table that looks something like this.
| Name | Sequence | Site Length | Overhang | Cut Positions | Frequency |
|---|---|---|---|---|---|
| PvuI | cgat/cg | 6 | three_prime | 44 | 1 |
| DpnI | ga/tc | 4 | blunt | 43 | 1 |
| HaeIII | gg/cc | 4 | blunt | 5 | 1 |
| BsaJI | c/cnngg | 4 | five_prime | 31, 57 | 2 |
| BssKI | /ccngg | 4 | five_prime | 31, 57 | 2 |
Each cut position indicates the base immediately 5' to the cut, e.g. 31 means that the enzyme cuts between nucleotides 31 and 32.
Each link in the Name column leads to the REBASE entry for that enzyme.
For further info about RestrictionMapper, check out the FAQ page. If that doesn't help, or if you have found a problem or a bug, please email Peter Blaiklock at webmaster |AT| restrictionmapper.org.
Virtual Digest Directions
Paste your sequence into the Paste Sequence Here box, select enzymes to digest with from the enzyme list and click Virtual Digest. Note that this function requires enzymes to be selected from the list. You will get a table of digestion products containing, for each product, the sequence, length, starting and ending positions in the original sequence and the 5' and 3' enzymes.
Note: Virtual Digest simulates simultaneous digestion. If your digestion conditions require sequential digestion, do a virtual digest with the first enzyme, then do virtual digests on the resulting fragments with the next enzyme, etc. This will ensure the most accurate picture of your final results.