Birgitte Binderup Simen - Orange CT, US Christine Lubeski - Niantic CT, US Jan Fredrik Simons - Burlingame CA, US
Assignee:
454 Life Sciences, A Roche Company - Branford CT
International Classification:
C12Q 1/68 C12P 19/34 C07H 21/04
US Classification:
435 612, 435 912, 536 2433
Abstract:
In one embodiment of the invention a method for detecting low frequency occurrence of one or more HIV sequence variants associated with drug resistance is describe that comprises generating cDNA species from RNA molecules in an HIV sample population; amplifying first amplicons from the cDNA species, wherein each amplicon comprises amplified copies and is amplified with a pair of nucleic acid primers that define a locus; clonally amplifying the amplified copies of the first amplicons to produce second amplicons that comprise an immobilized population of substantially identical copies from one of the amplified copies of first amplicons; determining a nucleic acid sequence composition from at least 100 of the immobilized populations in parallel on a single instrument; detecting one or more sequence variants that occur at a frequency of 5% or less in the nucleic acid sequence composition of the at least 100 immobilized populations; and correlating the detected sequence variants with variation associated with HIV drug resistance.