Open AccessA contaminant-free assessment of Endogenous Retroviral RNA in human plasma
Author(s) -
Timokratis Karamitros,
Dimitrios Paraskevis,
Angelos Hatzakis,
Mina Psichogiou,
Ioannis Elefsiniotis,
Tara Hurst,
Anna María Geretti,
Apostolos Beloukas,
John Frater,
Paul Klenerman,
Aris Katzourakis,
Gkikas Magiorkinis
Publication year - 2016
Publication title -
scientific reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.24
H-Index - 213
ISSN - 2045-2322
DOI - 10.1038/srep33598
Subject(s) - rna , endogenous retrovirus , genome , biology , contamination , computational biology , virology , human genome , gene , genetics , ecology
Endogenous retroviruses (ERVs) comprise 6–8% of the human genome. HERVs are silenced in most normal tissues, up-regulated in stem cells and in placenta but also in cancer and HIV-1 infection. Crucially, there are conflicting reports on detecting HERV RNA in non-cellular clinical samples such as plasma that suggest the study of HERV RNA can be daunting. Indeed, we find that the use of real-time PCR in a quality assured clinical laboratory setting can be sensitive to low-level proviral contamination. We developed a mathematical model for low-level contamination that allowed us to design a laboratory protocol and standard operating procedures for robust measurement of HERV RNA. We focus on one family, HERV-K HML-2 (HK2) that has been most recently active even though they invaded our ancestral genomes almost 30 millions ago. We extensively validated our experimental design on a model cell culture system showing high sensitivity and specificity, totally eliminating the proviral contamination. We then tested 236 plasma samples from patients infected with HIV-1, HCV or HBV and found them to be negative. The study of HERV RNA for human translational studies should be performed with extensively validated protocols and standard operating procedures to control the widespread low-level human DNA contamination.