Lebedeva T, Harrison K, Fresia B, Ohashi M, Yu N
HLA Laboratory, American Red Cross, Northeast Division, Dedham, MA 02026, USA.
Tissue Antigens. 2009 Oct;74(4):339-42. doi: 10.1111/j.1399-0039.2009.01326.x.
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 DQB10301 and *0319 and resolves some common trans-ambiguities.
该项目的目标是获取DQB1基因第3内含子的序列，以便开发一种基于测序的分型方案，该方案能够提供完整的DQB1基因第3外显子序列。目前的方案无法提供第3外显子的完整序列，因此无法区分常见且记录充分的等位基因DQB10301和0319，也无法使用位于641和650位置的组特异性测序引物（GSSP）测序来解决一些常见的跨模糊性问题。对最常见DQB1等位基因纯合的样本用于获取第3内含子序列，这些序列用于设计基于内含子的引物以扩增第3外显子。该方案经过了广泛验证；未观察到等位基因缺失。所提出的方案能够对DQB10301和0319进行鉴别分型，并解决一些常见的跨模糊性问题。
