SPTB基因c.5798+1G>A新变异导致遗传性球形红细胞增多症一家系  

Hereditary spherocytosis due to a novel c.5798+1G>A variant of the SPTB gene

作　　者：刘红彦 黄佳 姜迎海 郭梁洁 肖海 王红丹 Liu Hongyan;Huang Jia;Jiang Yinghai;Guo Liangjie;Xiao Hai;Wang Hongdan(Institute of Medical Genetics,Henan Provincial People’s Hospital of Zhengzhou University,People’s Hospital of Henan University,Zhengzhou,Henan 450003,China)

机构地区：[1]河南省人民医院医学遗传研究所(郑州大学人民医院),河南大学人民医院,郑州450003

出　　处：《中华医学遗传学杂志》2020年第1期17-20,共4页Chinese Journal of Medical Genetics

基　　金：河南省科技攻关计划项目(182102310170)。

摘　　要：目的对一个遗传性球形红细胞家系致病基因进行鉴定和突变致病性分析。方法采集该家系共17人的外周血样。采用高通量测序分析先证者外周血DNA,筛选候选致病变异并进行家系共分离分析。然后采用同源重组构建pCAS2c.5798+1G和pCAS2c.5798+1A型质粒,转染293T细胞,应用反转录PCR、TA克隆和Sanger测序分析候选致病变异对剪接的影响,同时提取患者外周血RNA,分析候选致病变异在体内的剪接情况。结果高通量测序结果显示先证者携带SPTB基因c.5798+1G>A变异,且该变异与家系患者的表型共分离。体、内外剪接实验结果显示c.5798+1G>A变异明显影响RNA的正常剪接,导致阅读框移码产生提前终止的密码子。结论SPTB基因c.5798+1G>A新变异是导致该家系遗传性球形红细胞症的原因。Objective To explore the genetic basis of a pedigree affected with hereditary spherocytosis.Methods Peripheral blood samples were collected from 17 members of the pedigree.Genomic DNA of the proband was subjected to next generation sequencing.Candidate variant was validated by co-segregation analysis.pCAS2c.5798+1G and pCAS2c.5798+1A plasmids were constructed by homologous recombination and transfected into 293T cells.Reverse transcription PCR,TA cloning and Sanger sequencing were used to analyze the effect of candidate variant on splicing.Meanwhile,peripheral blood RNAs were extracted to analyze the effect of candidate variant on splicing in vivo.Results The proband was found to carry a c.5798+1G>A variant of the SPTB gene.The variant has co-segregated with the phenotype in the pedigree.In vitro and in vivo splicing experiments confirmed that the mutation has significantly affected the splicing,resulting in shift of reading frame and produced a premature termination codon.Conclusion The novel c.5798+1G>A variant of the SPTB gene probably underlies the pathogenesis of hereditary spherocytosis in this pedigree.

关 键 词：SPTB基因 遗传性球形红细胞症 剪接位点 基因变异 高通量测序 

分 类 号：R5[医药卫生—内科学]