Best DNA Extraction approach with three different sample types?

[grace-ac]

Hi!

I ran an experiment at FHL. Wanting to understand the ecology of the bacteria V. pectenicida that causes sea star wasting in sunflower sea stars --> can it be decreased in water by eelgrass and/or mussels.

The hypotheses are:

1. Presence of eelgrass will decrease the amount of V. pectenicida in seawater over time.
2. Presence of mussels will decrease the amount of V. pectenicida in seawater over time.
3. Presence of mussels + eelgrass will decrease the amount of V. pectenicid in seawater over time more than either alone.

Sample types I have:

1. Filters from water o capture if V. pectenicida is in the water (n=72)
2. Swabs from eelgrass tissue (n=32) --> does V. pectenicida form a biofilm on eelgrass?
3. Frozen mussels at -80C (n=40) --> DNA from mussel tissue AND swab shells --> Does V. pectenicida get concentrated in mussel tissue? Does V. pectenicida form a biofilm on mussel shells?

CURRENT PROCESSING NOTES:

1. I have processed filters from a pilot study from last year using ZymoBIOMICS DNA MIniPrep Kit (D4300).
2. It was suggested to me to use two other kits for the other sample types -->

• Qiagen DNeasy PowerSoil Kit for SWABS
• Qiagen Blood and Tissue Kit for MUSSEL TISSUE

QUESTION

In order to compare between these different sample types --> should I be using the same kit (in this case ZymoBIOMICS DNA Miniprep (D4300)) for all sample types?

The other two kits were suggested to me by people who have done work getting DNA from swabs and mussel tissue- but not targeting V. pectenicida.

[sr320]

No, I would not use Zymo D4300 for everything solely for comparability. Use the best-supported kit for each sample type - but keep the kit constant sample type

[kubu4]

should I be using the same kit (in this case ZymoBIOMICS DNA Miniprep (D4300)) for all sample types?

I don't think this matters (unless your goal is to compare extraction techniques).

With that said, you may want to perform extractions of serial dilutions of V.pectenicida cultures to give you an idea of what the lower limits of extraction might be for each of the kits. This adds a lot of extra work, though.