广西地区β地中海贫血杂合子β-LCR-HS2多态性序列分析

Analysis of Polymorphic Sequence in β-LCR-HS2 of β-thalassemia Heterozygous in Guangxi

关键词:  β地中海贫血  位点控制区  ^Gγ珠蛋白  基因  多态性  胎儿血红蛋白

Key words:  β-thalassemia  locus control region  ^Gγ-globin  gene  polymorphism  fetal hemoglobin

摘要:

分别应用PCR产物直接测序法和PCR-限制性内切酶Xmn酶切法分析17例广西地区β地中海贫血杂合子的34条染色单体上β-LCR-HS₂核心区AT重复序列(AT)_xN_y(AT)_z多态性类型及8580、8598、9114位点碱基改变、^Gγ珠蛋白基因启动子-158位点(^Gγ-158)单核苷酸多态性(SNP),并观察它们与高HbFβ地中海贫血杂合子的关系。结果共检出9种(AT)_xN_y(AT)_z类型:(AT)₉N₁₂(AT)₁₁、(AT)₈N₁₂(AT)₁₁、(AT)₉N₁₂(AT)₁₀、(AT)₁₀N₁₂(AT)₁₁、(AT)₉N₁₃(AT)₁₁、(AT)₉N₁₄(AT)₉、(AT)₁₀N₁₂(AT)₁₀、(AT)₈N₁₂(AT)₁₄和(AT)₉N₁₄(AT)₁₀,其中前3种类型检出最多(73.5%)。β-LCR-HS₂的8580、8598、9114位点均未发现突变。HbF>3%的12例受检者有6例^Gγ-158基因型为C/T,6例为C/C;作为对照的5例HbF正常(<2%)者均为^Gγ-158(C/C)。(AT)₉N₁₂(AT)₁₀主要在6例HbF>3%且^Gγ-158(C/T)的个体中检出(5/6)。提示(AT)₉N₁₂(AT)₁₀与HbF升高有一定关系,而且与^Gγ-158(C→T)突变存在不平衡连锁倾向。

Abstract:

34 chromosomes of 17 β-thalassemia heterozygous in Guangxi were studied. DNA sequence in the core region of β-LCR-HS₂ including AT repeats[(AT)_xN_y(AT)_z] and three single nucleotide polymorphisms(SNPs) at 8580,8598 and 9114 sites were detected by PCR-direct nucleotide sequencing and,SNP at-158 site of ^Gγ gene promoter(^Gγ-158) was detected by XmnⅠ restriction enzyme digesting after PCR amplification.The relationship between these polymorphisms and the elevation of fetal hemoglobin(HbF) are determined. 9 kinds of (AT)_xN_y(AT)_z motif were detected in 34 chromosomes,namely (AT)₉N₁₂(AT)₁₁,(AT)₈N₁₂(AT)₁₁,(AT)₉N₁₂(AT)₁₀,(AT)₁₀N₁₂(AT)₁₁,(AT)₉N₁₃(AT)₁₁,(AT)₉N₁₄(AT)₉,(AT)₁₀N₁₂(AT)₁₀,(AT)₈N₁₂(AT)₁₄ and (AT)₉N₁₄(AT)₁₀(AT)₉N₁₂(AT)₁₁,(AT)₈N₁₂(AT)₁₁ and (AT)₉N₁₂(AT)₁₀ were the most three patterns(73.5%). No mutation was found at 8580,8598 and 9114 sites of β-LCR-HS₂.Among 12 cases with HbF>3%,6 carried genotypes of ^Gγ-158 (C/T),while the others carried ^Gγ-158 (C/C). In control group,the genotypes of ^Gγ-158 of all 5 cases with normal HbF levels (<2%) were C/C. In 6 patients characterized by HbF>3% and ^Gγ-158(C/T),most of them carried (AT)₉N₁₂(AT)₁₀ motifs(5/6). Our study had implied that (AT)₉N₁₂(AT)₁₀ motif was linked to the elevation of HbF in β-thalassemia heterozygous from Guangxi,and had linkage disequilibrium with ^Gγ-158(C→T) mutation.