Question

An F' plasmid from an exotic marine bacterium that can degrade petroleum when mated with an F^-E. coli that cannot degrade petroleum results in an E. coli F' that can degrade petroleum. After sequencing the F' plasmid and comparing it to the F sequence, you see that it has about 6000 base pairs of additional DNA. You suspect that there is a gene in that sequence that encodes a protein that degrades petroleum. Describe how you would use reverse genetics to identify the gene involved in petroleum degradation (note: the 6000 bp sequence may have more than 1 gene encoded on it). In your answer, tell me what is the first thing that you will look for in the sequence to help you identify the gene and what are two additional sequence features you will look for to increase your confidence that you are looking at an actual protein encoding sequence. Finally, describe how you will test any candidate sequences that you identify as an actual gene that makes a petroleum degrading protein.

Answer 1

In order to find the gne involved in the petroleum degradation, the reverse genetics will be used. The DNA sequence in the additional DNA present in the F' plasmid will be analysed using the protein which degrades the petroleum. After the identification of the DNA sequence, the gene could be recognized that is giving rise to the concerned protein. The codon is the first thing that should be looked for in the gene. The amino acids contained in the protein should correspond with the DNA codon. Furthermore, the DNA contains non-coding sequences in between the coding sequences, these sequences too should correspond with the protein.

The confirmation can be done by incorporating the identified gene into a plasmid and then, studying its function.