Hi Isla,

Based on your experiment, probably the target gene is conserved in these specific strains, but you cannot affirm that is present in all mtb strains. You can BLAST the gene/protein sequence to verify if this gene is present in other strains too.

To confirm if the gene is essential, you can perform a knock-out mutation (there are some methodologies in literature).

Thank you Leandro Toshio Kochi for clarifying :)

in order to make any claims you would need to provide the supporting evidence, otherwise its just your opinion.

I would suggest sequencing your PCR products as you begin building your library.

Additionally, as Leandro mentions above, you should also BLAST these sequences, You may discover your gene of interest to be a homolog with conserved regions.

hope this helps,

Cheers

Gene specific primers are designed to target conserved regions within genomes... Getting these primers to amplify in your clinical strains is an indication of the strains being related by either belonging to the same species or genus! However, you need to further go ahead and sequence the PCR products to confirm their relatedness and also BLAST the sequence in NCBI to probably identify any other related sequences that may give you the identity of your PCR amplicons.

Essential genes are either constitutively expressed i.e always present and the other alternative is for the gene to be induced to be expressed upon need( induced expression). An essential gene may be only expressed upon need and I think you can not conclude that because the gene is present in all clinical strains, then it is essential!

Hi. In addition to the information given by Sarah, are you doing the PCR in gDNA or cDNA? Because the gene can be in there (gDNA), but not expressed (no amplicon in cDNA), therefore, it won't be essential.