I am trying to isolate S. pyogenes RNA after adhesion to A549 cells. The protocol states lysis of the adhered bacteria from cell line by saponin/ trypsin treatment followed by total RNA extraction. However the initial cell pellet is very less. I feel the chemicals are masking the RNA leading to hindrance in RNA isolation. Kindly suggest on how to isolate the RNA.
If reduced yield of "bacterial RNA" is the main concern, it may be important to know- (i) how many bacteria were in the co-culture? (ii) what would be the use of "bacterial RNA" after isolation? If the RNA is to be used for gene expression analysis of bacterial cells, the physical separation of bacteria from the cells of the cell line could be under more mild conditions.
Sir, I start with 107 bacterial cells of which around 105 get adhered to the A549 cells. I want to study the gene expression of the bacteria after adherence to A549 cells.
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