Systematic analysis of swine leukocyte antigen-DRB1 nucleotide polymorphisms using genomic DNA-based high-resolution genotyping and identification of new alleles
Systematic analysis of swine leukocyte antigen-DRB1 nucleotide polymorphisms using genomic DNA-based high-resolution genotyping and identification of new alleles
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DOI：
10.1111/j.1399-0039.2011.01662.x
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发表时间：
2011-06-01
期刊：
影响因子：
--
通讯作者：
Park, C.
中科院分区：
文献类型：
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作者：
Thong, L. Minh;Choi, H.;Park, C.
为了实现猪白细胞抗原（SLA）基因的全面高分辨率基因分型，我们对SLA-DRB1和DRB1假基因的不同等位基因的内含子1、2和外显子2的核苷酸多态性进行了系统分析。我们从不同的等位基因中扩增和克隆了16个SLA-DRB1和DRB2内含子1和2的部分序列，并将其与报道的4个SLA-DRB假基因DRB2、3、4和5的序列进行了分析。结果显示，在sla - drb相关基因的内含子1和内含子2中存在极端的核苷酸变异，包括替换和缺失。在此基础上，我们通过基因组聚合酶链反应（PCR）和随后的直接测序建立了一种全面的SLA-DRB1基因分型方法。共对415只动物进行基因分型，鉴定出18个DRB1等位基因的67个等位基因组合。其中，SLA-DRB1*kn04和*kn05两个等位基因此前未见报道。本研究的SLA- drb1基因分型结果与我们之前研究的SLA- dqb1基因分型结果相结合，发现了10个SLA II类单倍型，其中3个以前未报道过。用7个不同猪品种进行群体分析，发现SLA-DRB1等位基因频率在不同品种之间存在差异。我们的研究结果将有助于需要测序水平的SLA-DRB1基因分型结果的生物学实验，以及进一步利用野外样本研究SLA-DRB1的完整遗传多样性。
In an attempt to enable comprehensive high-resolution genotyping of the swine leukocyte antigen (SLA) gene, we performed a systemic analysis of nucleotide polymorphisms at introns 1 and 2 and exon 2 from diverse alleles of SLA-DRB1 and DRB1 pseudogenes. We amplified and cloned 16 partial sequences of SLA-DRB1 and DRB2 introns 1 and 2 from different alleles, and analyzed them together with sequences of four reported SLA-DRB pseudogenes, DRB2, 3, 4, and 5. The results showed the presence of extreme nucleotide variations within introns 1 and 2 of SLA-DRB-related genes including substitutions and deletions. On the basis of these results, we developed a comprehensive genotyping method for SLA-DRB1 by genomic polymerase chain reaction (PCR) and subsequent direct sequencing. A total of 415 animals were genotyped and 67 allelic combinations from 18 DRB1 alleles were identified. Among them, two alleles, SLA-DRB1*kn04 and *kn05, were previously unreported. SLA-DRB1 genotyping results from this study combined with those of SLA-DQB1 from our previous study presented 10 SLA class II haplotypes, three of which were previously unreported. Population analysis using seven different pig breeds showed differences in the allele frequency of SLA-DRB1 among breeds. Our results should benefit biological experiments requiring sequence-level genotyping results of SLA-DRB1 and further study of the complete genetic diversity of SLA-DRB1 using field samples.