Large-scale evolutionary surveillance of the 2009 H1N1 influenza A virus using resequencing arrays

Lee, Charlie Wah Heng; Koh, Chee Wee; Chan, Yang Sun; Aw, Pauline Poh Kim; Loh, Kuan Hon; Han, Bing Ling; Thien, Pei Ling; Nai, Geraldine Yi Wen; Hibberd, Martin L.; Wong, Christopher W.; Sung, Wing-Kin

We try to recover the remaining query bases that have NHIP of type described in Figure 2d or 2e by analysing the substitution bias from their PM and MM probes in the forward and reverse strands separately. Similar to how a mutation is confirmed, we compute the likelihood ' f that the observed PM probe (representing the unsure base call) is indeed the true PM probe of the sample sequence given the hybridization intensity-based ordering of its MM probes in the forward strand. We also compute a similar likelihood ' r for the PM probe in the reverse strand. If the PM probes in both strands are complementary and ' f , ' r > 2, the query base results in a strong base call (represented by upper case base calls 'A', 'C', 'G' or 'T'). However, in many cases, the PM probes in both strands are not complementary due to non-specific hybridization of MM probes in one or both strands. For such query bases, we make base calls based on ' f and ' r : if ' f > ' r and ' f > 2, a base call with weak support (represented by lower case base calls 'a', 'c', 'g' or 't') is made from the PM probe in the forward strand. Else, if ' r > ' f and ' r > 2, a base call with weak support is made from the PM probe in the reverse strand. Otherwise, they are assigned an unknown 'N' call.