1) when looking at a gel, how can i tell if there is evidence of genetic...

1) when looking at a gel, how can i tell if there is evidence of genetic evidence ( if i am looking at the presence of tubulin and 355 promoter)

2) how can i tell if there is nonspecific amplification present in a gel

Expert Solution

1. When you run gel either protein gel (PAGE) or nucleotide gel (agarose gel), you should have a reference ladder in a well. For, protein gel, peptides of different known molecular weight and for nucleotide gel, nucleotide of different molecular known weight which will indicate the presence of protein or nucleotide of certain molecular weight.

For, example, you are running a PAGE to detect tubulin in your sample. As we know the molecule weight of tubulin, by comparing the moleculer weight of the ladder and the band pattern of the sample, you can say if tubulin is present or not. If you see a band around the molecular weight region of tubulin which is colinear with the ladder, you will say tubulin is present, if not then it is absent.

It is also same for the detection of promoter through agarose gel. If the molecular weight of the promoter fragment come at the expected molecular weight region, then you have correctly amplified the specific promoter region.

2. When you do PCR, using specific primers against a particular region of the genome, it should amplify that specific region only which will be almost similar in molecular weight and should be observed in the agarose gel as a single band. But, if you see more than one band or a smear on the gel, there must be non-specific amplification had taken place. You can also identify the presence of the desired fragment by calculating the fragment's molecular weight and comparing with the nucleotide ladder.

Hope, this will help. If you still have specific queries, can ask me in comments.

gladiator answered 3 months ago

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