Create an experiment where you would use both a Southern blot and a Northern blot to determine if a newly isolated bacterium contains and actively uses the same catabolic pathway as previously demonstrated

Question

What is the value in doing the Southern blot first? With results from these experiments, how would you conclude if the same pathway is active or not?
What will be an ideal response?

Answer 1

Answer: Probes would be designed off gene sequences from the previously demonstrated pathway, and the same DNA probes could be used for both the Southern and Northern blots. With isolated genomic DNA from the isolated bacterium, the probes would be applied to visualize the presence of each gene. All probes hybridizing to the genome suggests the pathway exists in the isolate, and the probes would then be used to determine is those genes were all active during the catabolism of the particular substrate. The substrate inducing the catabolic pathway would be provided to the isolate as a sole carbon source and RNA harvested to determine which transcripts are present. Activity of the genes for the pathway would be identified with probes again hybridizing to the same sequences as in the Southern. The Northern would not have been necessary to perform in all the genes were not first identified in the genome. To conclude the same genes are responsible for the catabolic pathway as previously documented, all gene/transcript probes would have to hybridize to the transcriptome.