What genes/markers are you wanting to target? Or anything that works as a molecular identifier?

Hello Asif! Thank you for taking time with my question.

Well, there are several markers that have been identified for my organisms.

Example:

Cryptosporidium parvum:

60kD glycoprotein gene (gp60)

Full-length SSu rRNA (1733-1750 bp)

Cyckospora cayetanensis:

70 kD Heat shock protein (HSP70)

But these are specific primers up to the species level...

Since my samples are water and sediments, the idea is if there is the presence of the target organisms there, then one should expect multi species of that genus (Cryptosporidium spp. or Cyclopspora spp.).

If I will be using specific primers then I would be able (if present) identify Cryptosporidium parvum and Cyclospora cayetanensis only.

So in effect, other cryptosporidium spp. and other Cyclopspora spp. will be left out.

More so, if I use these specific primers only, then there is a big possible that the results would come out negative for the parvum and cayetanensis species but the samples may be have other species that cannot be detected by the specific primer set used.

So in effect, the use of specific primers enhances the specificity of the researchers result but the trade off is there is a big chance of not identfying other species present.

On the other hand, using non specific primers (18S rRNA) primers would be able to catch a broader spectrum of eukaryotic organisms but how would the researcher then resolve each species contained therein the sample.

I am a newbie in molecular techniques and have a difficult time understanding the protocols I am reading from my RRLs.

Which brings us back to my concern.

Is it possible for us to be able to use a primer which can catch all Cryptosporidium species and a primer which will catch all Cyclospora species? how do we do these?

Your suggestions would be most valuable for the progress of the molecular part of my research and is deeply appreciated!

Can you suggest any protocol, paper, or primer sets to identify these organisms at a genus level? and later on resolve the PCR products for species identification?