Blotting and Operon

[Transformers]

I had multiple choice question....how would you determine that the structure of a DNA sequence (w/3 adjacent genes) was in fact an operon using probes and blotting?

a. Southern blotting revealed 3 bands
b. Southern blotting revealed 1 band
c. Northern blotting revealed 3 bands
d. Northern blotting revealed 1 band

I put b because the DNA segment is 1 band...is this right guys?

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[unique135]

It doesn't make sense to use Southern Blotting. It would only detect the presence of DNA.

Since it is operon, the translation must be bacterial and polycistronic. Polycistronic mRNA, basically, carries multiple genes and would contain operon sequences. In Northern blotting, probes would bind to this single mRNA and thus one probes-mRNA band.

D should be the answer.

 

[gunj122]

It doesn't make sense to use Southern Blotting. It would only detect the presence of DNA.

Since it is operon, the translation must be bacterial and polycistronic. Polycistronic mRNA, basically, carries multiple genes and would contain operon sequences. In Northern blotting, probes would bind to this single mRNA and thus one probes-mRNA band.

D should be the answer.

wait, how does mRNA have operon sequences? an operon is based off of genomic DNA, not mRNA. am i missing something here?

shouldn't the answer be A? southern blotting, with the use of the right restriction enzymes, would reveal 3 different bands for the promoter, operator, and structural gene.

 

[Catburr]

wait, how does mRNA have operon sequences? an operon is based off of genomic DNA, not mRNA. am i missing something here?

shouldn't the answer be A? southern blotting, with the use of the right restriction enzymes, would reveal 3 different bands for the promoter, operator, and structural gene.

You aren't missing anything. It should be A. You want to check a DNA sequence, so it's a Southern blot. Verification based only on the size of the whole sequence (1 band) would be pretty crummy verification. Yeah, you'd attempt to cut it up (multiple bands), and if it cut into the sizes you predicted based on the sequence you expect to be present, you're good.

 

[whiteshadodw]

It doesn't make sense to use Southern Blotting. It would only detect the presence of DNA.

Since it is operon, the translation must be bacterial and polycistronic. Polycistronic mRNA, basically, carries multiple genes and would contain operon sequences. In Northern blotting, probes would bind to this single mRNA and thus one probes-mRNA band.

D should be the answer.

gonna have to agree here. at first i thought you'd want to check for DNA. however, if the operon is in fact an operon, then you'd expect the DNA to be transcribed into RNA containing all 3 genes. you can cut up the mRNA and use 3 separate probes to verify that the mRNA that was translated did in fact have the 3 genes. its possible that it could have only 1 gene or 2 and that the 3rd gene is part of another operon or something else.

an alternative but similar experiment would have changes to the promoter in the operon. the promoter sequence that gives the most mRNA would be the 'best' or rather consensus promoter sequence.