Hi, as i want to extract bacterial DNA from mice tissue, but the final solution also contains both bacterial DNA and background DNA (mice tissue DNA), Will the somatic DNA influence the bacterial DNA when doing downstream PCR analysis? i mean even if i use the bacterial specific primer, will the somatic DNA influence the PCR results?? if so, how to reduce this influence? many thanks!
I think if you are using bacteria specific primers then mice DNA will not affect your PCR reaction.
or you could go for Decontamination procedures for your Bacteria,
or you could first culture your bacteria then go for DNA isolation(if possible).
All the best
Decontamination here means removal of unwanted DNA... it just enrichment of your desired microbe. by removing other microbes. for example in Mycobacterium we use NALC-NaOH method to increase recovery of mycobacterium .
Why don't you use mice dna extract as negative control in your PCR? It would inform you on the specificity. Do you have problem in the PCR yield ?
I would try PCR directly but if you experience any sensitivity issues, you can enrich bacterial DNA in several ways:
with NebNext (https://www.neb.com/products/e2612-nebnext-microbiome-dna-enrichment-kit)
with Molysis (http://www.molzym.com/index.php/products/dna-isolation-products/pathogen-dna-molysis) - I dont kow if that works for the mice tissue you are using, or whether it is possible to adapt the kit to your needs, but if you contact them they can certainly help
or using nuclease suppression (http://www.sciencedirect.com/science/article/pii/S0003269711007421, http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0103610)
- Badges
- Science method