In previous articles, I have discussed the structure and function of certain proteins, such as adrenaline receptors, ATP synthase and those involved in vesicular transport. In this case, it was proteins and the cardinal point assigned to them was the west. A clever twist: western blotting techniqueĪfter the sensation created by those two techniques, another biomolecule was added to the increasing repertoire of gel probing experiments. I recommend watching this video if you are interested in learning more about how the two techniques mentioned above are carried out. The experimental method is almost identical, with the exception of the nucleotides being used (ribonucleotides instead of deoxyribonucleotides). Therefore, if a cell contains more or less mRNA for a specific gene than a healthy cell, this may be caused by a disease, since there is more or less of that protein being synthesised. The difference with respect to Southern blotting is that RNA is dependent on gene expression. Northern blotting allows researchers to find which RNA fragments are present in cells at a given time. Of course, they could not come up with a better name for it than northern blotting. Instead of looking for DNA sequences, the new method probed RNA sequences. Shortly after the Southern technique was published in 1975, another group of scientists came up with an analogous analysis. If you were waiting for the punchline of the joke, here it is. The witty joke: northern blotting technique To carry out Southern blotting, the DNA must be denatured so that the probe can attach to one strand of the gene by base pairing complementarity. If Sodium Dodecyl Sulfate (SDS) solution is applied to the proteins before pipetting them onto the gel, they will denature and move along the gel, like DNA. Gel electrophoresis can also be used to separate proteins. Below is a visual representation of gel electrophoresis. Smaller molecules will be able to move more rapidly through the pores, while larger molecules will get tangled in the meshwork and take longer to traverse the gel. The reason why the separation occurs on the basis of size is because the gel is a meshwork. In the case of DNA where the phosphate backbone is negatively charged, it moves from the cathode (negative) to the anode (positive) of the tank in which the gel is placed. As the name indicates, a sample is placed on a gel, which, when subjected to an electrical current induces the movement of the molecules in the sample. The objective of a gel electrophoresis is to separate molecules according to size. If you are already familiar with this procedure, you can skip to the next section. Gel electrophoresisĪs an aside, I am going to explain in a bit more detail what a gel electrophoresis is and how it works. Once you image the nitrocellulose membrane, only those DNA fragments which were complementary to the probe that was added will be visible.
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