User:Anthony Salvagno/Notebook/Research/2009/01/26/Restriction Endonucleases

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I tend to go here for the most basic information and since it always seems like a good starting point. After I read about them there I will check out the link Koch provided me (next section).Template:Ks

Nothing too complicated here. Rest. Enzymes, cut the DNA at a specific location. The site can either be palindromic (where the complimentary strand is the inverse of the other) or not. You can also have sticky ends or blunt endsSJK 12:33, 28 January 2009 (EST)
12:33, 28 January 2009 (EST)
T4 DNA ligase can link blunt ends, but the efficiency is lower. It's also trickier to prevent circularization of fragments, if your fragments have two blunt ends. Although you can due things by removing phosphate groups from one piece with CIAP (calf intestinal alkaline phosphatase). For us, I'm guessing specific sticky ends are going to be very helpful for us, and I don't have any specific blunt-ended procedures in mind
. The sticky end allows for a complimentary strand to be attached (via ligation) to the site. I don't know what you can do with blunt ends.

Wikipedia goes into the types of RE's and from the looks of it Type II one's are the most used. Unfortunately the naming of these things are not the same as the type (ie EcoRI is not a type I RE).

Restriction Enzymes Overview

I don't like this page... the font hurts my eyes.

Interesting note:

Type I enzymes are complex, multisubunit, combination restriction-and-modification enzymes that cut DNA at random far from their recognition sequences. Originally thought to be rare, we now know from the analysis of sequenced genomes that they are common. Type I enzymes are of considerable biochemical interest but they have little practical value since they do not produce discrete restriction fragments or distinct gel-banding patterns.

Maybe we can use these for SDM since they may provide random fragments of unknown size. It seems the cut locations are also random and so the locations too would not be known. Another sort of proof-of-principle.

SJK 12:35, 28 January 2009 (EST)
12:35, 28 January 2009 (EST)
I agree that they're fascinating! There may be in vitro assays you could do to study them by unzipping, since they probably (OK, "maybe") form loop structures that you could detect.

Some words to learn:

  • homodimers
  • heterodimers
  • monomers