Hi,
I’m currently working on phages against pathogenic E.coli and I have come across this article of rapid identification of selected phages through Multiplex PCR. This method allows detection at very initial stage and whole genome sequencing of phage is still required to confirm preliminary classification. Should I go for Multiplex PCR first or directly shift to whole genome sequencing?
There are several methods to identify phages including 16S rDNA sequence analysis was used to identify the host bacterium, electron microscopy, one-step growth curve, stability studies, the whole genome sequencing and assembly, identification of the phage receptor and phage inactivation by LPS. Multiplex PCR detects in one reaction the presence of phages in samples, so I think you should go straight for it, except there are other information that you are interested in. Article Multiplex PCR for Detection and Identification of Lactococca...
Hi,
there are different approaches which in the end result in different qualtities of data output. On the other site keep in mind, that the demand in regard of sample amount and purity might differ e.g. some Multiplex mixes are inhibitor resistant and can amplify samples which woudl not be suitable for shotgun sequencing. Let me know if you need more input regarding the mixes
Sven
Hi,
I think the multiplex PCR described in the paper is wort to try. In addition, you can try to check the genomic patter of your phages using Restriction Fragment Length Polymorphism (RFLP) by extracting the phage genome and digest it using various restriction enzyme.
If your purpose is to get broad host range phage, I would check (1) the host range, (2) RFLP or multiplex PCR, then (3) whole genome sequence using the best candidate phage.
Good luck!
Multiplex pcr for phages is not that worth as in bacteria because it lacks uniformity in conserved genes.Go for WGS and TEM
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