2.5.3 Tests for Selection
For selection, we conduct a priori classification of peptide binding
region (PBR) and non- peptide region upon inferred passerine PBR
sequences [40, 41] homology sites with chicken MHC [42, 43] and
human HLA [44]. The identification of sites subjected to selection
in MHC class I Exon 3 was performed using various methods. The first
standard selection test (Tajima’s D, Fu & Li’s F* and Fu & Li’s D*)
were calculated using DnaSP 5.0 [32]. Second method was the
calculation of parameter (ω) for functional alleles. It was carried out
an overall estimation ofd N/d S of MHC class I Exon
3 and the other was codons comprising only PBR and non-PBR, which was
calculated with MEGA 7.0 according to the Nei-Gojobori method [45]
with the Jukes and Cantor correction. Standard error estimates were
derived from 1000 bootstrap replicates. Z test of historical positive
selection [46] was calculated in MEGA 7.0. Third, the Maximum
likelihood implemented in codeml in PAML 4.9 was used for identification
of sites involved in the positive selection, which are indicated where
the ratio ω
(d N/dS ) larger than 1
[47]. Two different models corresponding ω were tested: M7 (beta),
M8 (β and ω). To find whether the
alternative model (M8) provided better fitter than the M7, we performed
Likelihood ratio tests to compare twice the difference of the
log-likelihood ratios (2ΔlnL) using a distributionχ 2. PSSs in the M8 model was identified by PP
more than 95% using the Bayes empirical Bayes procedure. Positively
selected sites were verified at each codon site separately using many
complementary approaches implemented in datamonkey
(http://www.datamonkey.org/) [48] in addition to afore mention
methods. Specifically, we used MEME [49], FEL, SALC [50] and
FUBER [51]