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Assessment of differences between DNA content of cell-cultured and freely suspended oocysts of Cryptosporidium parvum and their suitability as DNA standards in qPCR

Woolsey, Ian D.; Blomstrand, Berit Marie; Øines, Øivind and Enemark, Heidi L. (2019) Assessment of differences between DNA content of cell-cultured and freely suspended oocysts of Cryptosporidium parvum and their suitability as DNA standards in qPCR. Parasites & Vectors, 12 (596), pp. 1-17.

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Online at: https://parasitesandvectors.biomedcentral.com/articles/10.1186/s13071-019-3851-7

Summary

For many studies involving Cryptosporidium, only relative DNA content is required and as such, the superior linearity afforded by freely suspended oocysts and diluted DNA template (from either cell culture derived standards or freely suspended oocysts) will allow for more accurate relative quantification in each assay. Parasite division in the cell culture standards likely explains the higher DNA content found. These standards, therefore, have the potential to more accurately reflect DNA content in cell culture assays, and despite more modern methods available for absolute quantification, i.e. droplet digital PCR (ddPCR), the ubiquity of qPCR for the foreseeable future encourages further investigation into the reduced linearity observed in these standards such as varying oocyst seeding density, non-linear growth rates and assay efficiency.


EPrint Type:Journal paper
Keywords:Cryptosporidium, HCT-8, Cell culture, qPCR, Barkcure,
Subjects: Animal husbandry
Animal husbandry > Health and welfare
Research affiliation: Norway > NVI - National Veterinary Institute
Norway > NORSØK - Norwegian Centre for Organic Agriculture
ISSN:1756-3305
DOI:10.1186/s13071-019-3851-7
Deposited By: Land, Anita
ID Code:37180
Deposited On:03 Feb 2020 07:46
Last Modified:03 Feb 2020 07:46
Document Language:English
Status:Published
Refereed:Peer-reviewed and accepted

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