DQB1 intron 3 sequences provide an insight into the evolution of DQB1 alleles and form the basis for intron-based sequencing

The goal of this project was to obtain sequences of intron 3 of DQB1 in order to develop a sequencing-based typing protocol that provides a complete DQB1 exon 3 sequence. Current protocols do not provide complete sequences of exon 3, thus not allowing to differentiate common and well-documented alleles DQB1*0301 and *0319 and resolve some common trans-ambiguities with group-specific sequencing primer (GSSP) sequencing using positions 641 and 650. Samples homozygous for the most common DQB1 alleles were used to obtain intron 3 sequences, which were used to design intron-based primers for exon 3 amplification. The protocol was extensively validated; no allele dropouts were observed. The presented protocol allows differential typing of DQB1*0301 and *0319 and resolves some common trans-ambiguities.