The specific sites under positive selection according to the models 2a and 8 were identified by a Bayesian approach.
In addition we conducted dN/dS calculations using the method by [ 50] implemented in PAML [ 87] for the 122 putative genes under positive selection according to dR/dC.
As for gene loss, it may also cause positive selection according to the dosage-compensation model.
6 Number of codons under positive selection according to TreeSAAP [ 35]. 7 Number of codons under positive selection changed on the branch of the group A-B boundary according to TreeSAAP.
Total CD4+ T cells were purified from spleen and peripheral lymph nodes (LNs) by positive selection using immunomagnetic beads according to the manufacturer's protocol (EasySep Biotin Selection Kit, StemCell Technologies Inc ,Vancouver, BC, CA).
CD4+ T cells were isolated by positive selection using CD4 beads, according to protocols provided by the manufacturer (Miltenyi, Bergisch Gladbach, Germany; purity was generally higher than 95%), and cultured in human T cell culture medium (Lonza, Walkersville, MD, USA).
PBMCs were isolated by Ficoll-Hypaque density gradient centrifugation (Hengxin Chemical Reagent Co, Ltd., Shanghai, China) and CD4+ T cells and B cells were isolated by positive selection using magnetic beads according to the manufacturer's protocol (Miltenyi Biotec, Bergisch Gladbach, Germany).
2D2 CD4+ T cells were isolated from spleen and LN by positive selection using MACS (Miltenyi Biotech) according to the manufacturer's instructions.
Cells isolated from spleen cell preparations were labeled with anti-CD4 micro-beads and separated by positive selection on a magnetic column, according to the manufacturer's instructions (Miltenyi Biotech).
Blood samples (30 ml) from fasting subjects were collected into EDTA, and monocytes were isolated by positive selection with CD14 microbeads (Miltenyi) according to the manufacturer's instructions.
