Liu Li, Muralidhar Shalini, Singh Manisha, Sylvan Caprice, Kalra Inderdeep S, Quinn Charles T, Onyekwere Onyinye C, Pace Betty S
Department of Molecular and Cell Biology, Sickle Cell Disease Research Center, University of Texas at Dallas, Richardson, TX 75083, USA.
Blood Cells Mol Dis. 2009 Jan-Feb;42(1):16-24. doi: 10.1016/j.bcmd.2008.07.002. Epub 2008 Oct 1.
Five major beta-globin locus haplotypes have been established in individuals with sickle cell disease (SCD) from the Benin, Bantu, Senegal, Cameroon, and Arab-Indian populations. Historically, beta-haplotypes were established using restriction fragment length polymorphism (RFLP) analysis across the beta-locus, which consists of five functional beta-like globin genes located on chromosome 11. Previous attempts to correlate these haplotypes as robust predictors of clinical phenotypes observed in SCD have not been successful. We speculate that the coverage and distribution of the RFLP sites located proximal to or within the globin genes are not sufficiently dense to accurately reflect the complexity of this region. To test our hypothesis, we performed RFLP analysis and high-density single nucleotide polymorphism (SNP) genotyping across the beta-locus using DNA samples from healthy African Americans with either normal hemoglobin A (HbAA) or individuals with homozygous SS (HbSS) disease. Using the genotyping data from 88 SNPs and Haploview analysis, we generated a greater number of haplotypes than that observed with RFLP analysis alone. Furthermore, a unique pattern of long-range linkage disequilibrium between the locus control region and the beta-like globin genes was observed in the HbSS group. Interestingly, we observed multiple SNPs within the HindIII restriction site located in the Ggamma-globin intervening sequence II which produced the same RFLP pattern. These findings illustrated the inability of RFLP analysis to decipher the complexity of sequence variations that impacts genomic structure in this region. Our data suggest that high-density SNP mapping may be required to accurately define beta-haplotypes that correlate with the different clinical phenotypes observed in SCD.
在来自贝宁、班图、塞内加尔、喀麦隆和阿拉伯 - 印度人群的镰状细胞病（SCD）患者中，已确定了五种主要的β - 珠蛋白基因座单倍型。历史上，β - 单倍型是通过对β - 基因座进行限制性片段长度多态性（RFLP）分析来确定的，该基因座由位于11号染色体上的五个功能性β - 样珠蛋白基因组成。此前试图将这些单倍型作为SCD中观察到的临床表型的可靠预测指标，但均未成功。我们推测，位于珠蛋白基因近端或内部的RFLP位点的覆盖范围和分布不够密集，无法准确反映该区域的复杂性。为了验证我们的假设，我们使用来自正常血红蛋白A（HbAA）的健康非裔美国人或纯合子SS（HbSS）疾病患者的DNA样本，对β - 基因座进行了RFLP分析和高密度单核苷酸多态性（SNP）基因分型。利用88个SNP的基因分型数据和Haploview分析，我们生成的单倍型数量比仅通过RFLP分析观察到的更多。此外，在HbSS组中观察到基因座控制区与β - 样珠蛋白基因之间存在独特的长程连锁不平衡模式。有趣的是，我们在位于Gγ - 珠蛋白内含子序列II中的HindIII限制性位点内观察到多个SNP，它们产生相同的RFLP模式。这些发现表明RFLP分析无法解读影响该区域基因组结构的序列变异的复杂性。我们的数据表明，可能需要进行高密度SNP图谱分析，以准确确定与SCD中观察到的不同临床表型相关的β - 单倍型。
