Využití deep-sequencing při studiu závislostí průběhu lentivirové infekce na variabilitě receptorů přirozené imunity.

• Main Author: Zelníková, Jana
• Other Authors: Jílek, Petr
• Format: Dissertation
• Language: Czech
• Published: 2014
• Online Access: http://www.nusl.cz/ntk/nusl-336627

Jana Zelníková Analysis of association of MHC and KIR variants with the course of lentivirus induced disease by next-generation sequencing Thesis Univerzita Karlova v Praze, Farmaceutická fakulta v Hradci Králové Farmacie Background: The aim of this work was to use the SIV infection in macaques (the only available model of human HIV infection) to show potential differences in MHC I haplotypes in macaques experimentally infected with SIVmac239 (or SHIV clade C) and their correlation with low or high viral loads, using pyrosequencing. Methods: The RNA samples from leukocytes from experimental macaques were reversely transcribed into cDNA and subjected to PCR of the 190 bp fragment of MHC I using fusion primers containing MIDs and FastStart High Fidelity Taq DNA polymerase. Subsequently the PCR generated samples were subjected to pyrosequencing using GS Junior System (Roche) according to the manufacturer`s protocol. The generated data were analyzed by a series of data filtration steps using scripts in Python and subsequently by DNAStar Lasergene 11. Final alleles were identified by ncbi-blast+ 2.25. Results: A total of 98 known aleles and 11 new aleles of MHC class I were found. The analysis identified those aleles, which correlate with fast or slow disease course. The aleles associated with slow...