Our recent study demonstrated that defects in p110δ result in B‐cell immunodeficiency that is very similar to that observed in BTK‐deficient mice. We revealed that the p110δ fit the B‐cell signal transduction complex and played a non‐redundant role in the development and function of B cells. In humans, most children with primary B‐cell immunodeficiency have mutations in the BTK, whereas a few have defects in the components of the B‐cell signal transduction complex. But little is known about the genetic variation of p110δ in children with defects in B‐cell immunodeficiency of unknown aetiology. Sixteen patients from 15 unrelated families and 112 normal controls underwent sequence analysis to identify genetic variations of the p110δ. Allele frequency in each group was also analysed and compared. We identified five single base‐pair polymorphic nucleotide exchanges in both patient and control groups with similar allele frequencies, which did not contribute to the immunodeficiency. Three of them are novel (m.953A>G, m.1200C>T and m.1561A>G), and the m.953A>G and m.1561A>G nucleotide exchanges are non‐synonymous (N253S and T456A, respectively). The novel m.1561A>G was in complete linkage disequilibrium with the known m.873A>G in our study of Taiwanese group. In addition, one novel single base‐pair missense mutation, m.3256G>A (E1021K), was identified in one boy with typical clinical features of primary B‐cell immunodeficiency and could not be found in either his family or the normal control population. By atomic structural analysis of the amino acid as well as the alignment comparison between species, it resulted in the replacement of the negative‐charged amino acid E with the positive‐charged amino acid K at codon 1021, located in the highly conservative and important catalytic functional domain. Our findings could shed light on further understanding the polymorphisms of p110δ in B‐cell immunodeficiency and different populations. Moreover, the 3256G>A missense mutation raised the attention and warranted further extensive analysis to elucidate the role of p110δ in human immunodeficiency.