Skip to main content

Cre lox system - Basics

It's been a long time, since I wrote a post here. Finally, final year of my course and so very busy :) with books! And now, got time to share with you.

As you know already (don't know?, then get to know :P ), I use to have favorite subjects in each semester, third semester it was microbiology, then in fourth semester, it was cell biology, fifth was molecular biology, sixth was genetic engineering and now i'm in seventh semester, but, you know what, I don't have a favorite subject!!! Don't worry that I lost interest, instead, I got 3 favorite subjects - Immunology, Animal biotechnology and Plant Biotechnology!

It's awesome this time to have more than one favorite subject and here I'm gonna tell you something about, cre-lox system which is most widely used for making modifications (mostly deletions). These modifications can be done at a specific tissue alone by using a tissue specific promoter i.e., you can selectively knock out a particular gene in particular cells (for eg: hepatocytes) alone.

Cre recombinase is a protein which could bind with specific sequences called loxP sequences. To explain it in a simple way, the cre recombinase binds at loxP sites on either side of your gene (which is to be deleted) and makes a cut! This leads to removal of the gene and further the break is repaired using the DNA ligase of host system.

A single loxP site will have 13bp on either side of middle 8bp. The cre recombinase protein binds with this 13bp sequences forming dimer i.e., one loxP site contributes for a dimer (containing 2 cre recombinase units). Similarly, loxP on the other side of the target gene will contribute for a dimer formation. Then these two dimers will form tetramer (by folding the DNA strand as shown in figure. Then, cutting at the tetramer site happens leading to either translocation, inversion or deletion depending on the direction of the loxP site (i.e whether it is direct or invert repeat on either side of the target gene or located on different chromosomes)

                         13bp                          8bp                  13bp
             ATAACTTCGTATA -NNNTANNN-TATACGAAGTTAT





This insertion, deletion and translocation could be done specifically at a particulat type of cell by using specific promoters. For example: To delete a particular gene in kidney cells alone, one could use kidney specific promoter for controlling the production of cre recombinase, so that cre recombinase will be expressed only in kidney cells. The molecular mechanisms of this deletions and recombinations invloves holliday junction formation. But, to be frank, i'm not clear or good at this holliday junction.

I''m trying to understand the molecular mechanism behind this, and, once I'm clear with it, I'll share it with you.

Understood the cre-loxp system? Got  any doubt? anything wrong in my explanation? Kindly comment and I would try to reply or correct as soon as possible.


Comments

  1. Lovely Your Post..............

    CRE Recombinase- GenTarget Inc - CRE Recombinase, from bacteriophage P1, catalyzes recombination between 34 base-pair target sequences, called lox sites. Purified CRE enzyme can join individual plasmids containing lox sites.

    ReplyDelete
  2. Nicely explained. I am also writing a blogs on life sciences topics. Kindly go through them and give me some suggestions regarding my blogs.

    http://netweget.blogspot.in/

    ReplyDelete

Post a Comment

Popular posts from this blog

Sea Butterfly!

Neih hou!  Don't roll you eyes wondering what it is! "Neih hou" is how you say "hello" in Cantonese. Guess what, I am in Hong Kong and therefore the language Cantonese. I came to Hong Kong this summer as an intern before officially joining as a PhD Student. This is my second experience abroad, far away from home, new language, new culture, I expected me to have a "really" bad cultural shock, but, actually I experienced only 50% of what I expected. For a girl who have used the marina beach only for eating "sundal", bikini in beach was a shock (FEEL FREE TO JUDGE ME!). The first time, I went to the big wave beach here, I was the only one who was totally covered, when everyone was enjoying their Beer, I was slowly sipping my lemon juice (THE ODD ONE OUT!). I realized how beautifully different the world outside is! There is no single standard for right or wrong, it varies in different countries, in different regions around the world. And it

The Butterfly flew for the first time!

Hi dear sweet lovely reader, Didn't meet you for a long time... Just after complaining about the stipend problem in the last post, I actually wanted to write a lot here, but, couldn't find time (Such a lazy girl, I am!). As you might be knowing, the author (me) is the butterfly here :P And, I flew for the first time!! YEAH! I FLEW! I flew like a butterfly! It was an awesome experience flying for the very first time! (ow, no! I dint put a trans-gene in my body and fly after getting  a pair of wings, but with a passport and VISA). I'm this girl, full of dreams wishing to do "this", "that" and "all kind of stuffs that one can do". And, one among those dreams was to fly one day! TO FLY FREE!(I mean, not free of cost :P, but liberation) I always wanted to fly in those big, big airplanes, but, I always have avoided that mode of transport. Reason? Obviously, it would burn a big hole in my dad's pocket (actually he can afford if I wan

Lowry Assay Principle and procedure

Though there are several protein assays available, the most preferred one in many laboratories is "Lowry assay". It is effective in the concentration range of 0.01 mg/ml to 1 mg/ml. And, as an additional info, the paper published describing the procedure and principle of Lowry Assay is the most cited paper in the scientific history. (Feeling like, "Wow! I want to publish one to compete with Oliver.H. Lowry"???  :P) Why Lowry?   Though there are several other protein assays, mostly Lowry assay is used in many laboratories. The reasons for preferring Lowry are: sensitivity of the assay, highly reproducible, cost effective, easy to perform. Biuret assay is generally used for higher protein concentrations like tissue samples but, Lowry for less concentrated samples and hence used in most of the molecular biology laboratories where there will be need for assaying comparatively less concentrated protein samples (in most cases where we attempt to produce enzymes). Oth