目的通过对1例新生儿期特殊面容、神经系统结构畸形患儿进行全外显子组序列检测分析,旨在为该患儿寻找潜在的致病原因。方法纳入1例在复旦大学附属儿科医院(我院)新生儿病房住院期间未能明确诊断的多发畸形患儿,主要临床表型为前额突出...目的通过对1例新生儿期特殊面容、神经系统结构畸形患儿进行全外显子组序列检测分析,旨在为该患儿寻找潜在的致病原因。方法纳入1例在复旦大学附属儿科医院(我院)新生儿病房住院期间未能明确诊断的多发畸形患儿,主要临床表型为前额突出、腭弓高、耳位低、枕部较平,双侧脑室扩大、透明隔部分缺如、胼胝体发育异常,采用Sure Selct Human All Exon捕获试剂盒和Illumina Hi Seq2000测序平台,行全外显子组序列检测。数据分析采用复旦大学附属儿科医院转化中心所建立的高通量测序数据分析流程。采用Sanger测序进行验证。结果患儿全外显子组序列检测数据,共检测到79 064个变异,经过质量控制筛选、变异频率筛选、变异分类筛选,剩余645个变异。在进一步分析中,645个变异中有159个其所在基因在OMIM数据库及HGMD数据库与疾病相关。从3个已经报道的突变位点中锁定致病突变为FGFR2基因(NM_000141)c.C1040G,p.S347C。Sanger测序在家系内验证该位点为新发(de novo)突变。结论采用全外显子组序列检测,明确诊断FGFR2相关疾病1例。并且结合我院已经建立的高通量测序数据分析和临床诊断流程,为新生儿多发畸形寻找潜在的致病基因提供了快速、高效的方法。展开更多
目的通过对1例智力发育迟缓患儿全外显子组测序(WES),旨在为该患儿寻找潜在的致病原因。方法纳入1例在复旦大学附属儿科医院(我院)住院期间诊断不明患儿,采用Sure Selct Human All Exon捕获试剂盒和Illumina Hi Seq2000测序平台,行WES检...目的通过对1例智力发育迟缓患儿全外显子组测序(WES),旨在为该患儿寻找潜在的致病原因。方法纳入1例在复旦大学附属儿科医院(我院)住院期间诊断不明患儿,采用Sure Selct Human All Exon捕获试剂盒和Illumina Hi Seq2000测序平台,行WES检测;数据分析采用我院分子诊断中心建立的高通量测序数据分析流程;结果采用Sanger直接测序法进行验证。建立体外大鼠神经元原代培养和糖氧剥夺(OGD)模型,分为空白对照组(Sham+PBS亚组、Sham+POSTN亚组)、OGD组(OGD+PBS亚组和OGD+POSTN亚组),采用免疫荧光、LDH毒性检测、Brd U和TUNEL方法检测POSTN对2组及其亚组神经元生长增殖和凋亡的影响。结果患儿WES共检测到532 151个变异,筛选后检测到POSTN基因(NM_006475)外显子23:c.A2475T:p.G825G和外显子19:c.G2251A:p.E751K。Sanger测序显示p.G825G来自母亲,p.E751K来自父亲,符合复合杂合遗传模式。培养的大鼠原代神经元(MAP-2或Neu N阳性细胞)中有内源性POSTN的表达。LDH活性Sham+POSTN亚组与Sham+PBS亚组差异无统计学意义,OGD+POSTN亚组与OGD+PBS亚组差异有统计学意义。OGD+PBS亚组致神经元细胞缺氧2、6、12 d后比Sham+PBS亚组Brd U阳性的神经元数量增高,TUNEL阳性神经元凋亡也增高;OGD+POSTN亚组各时间点神经元增殖较OGD+PBS亚组均明显增高,但凋亡减少。结论 POSTN可以减少脑损伤后LDH对于神经元的细胞毒性,促进损伤后神经元的增殖,抑制凋亡。展开更多
Major histocompatibility complex （MHC） is a family of highly polymorphic genes activating adaptive immunity in vertebrates. However, the underlying mecha- nism of MHC evolution is still not fully understood. In this...Major histocompatibility complex （MHC） is a family of highly polymorphic genes activating adaptive immunity in vertebrates. However, the underlying mecha- nism of MHC evolution is still not fully understood. In this study, we investigated genetic variation of three classical MHC class I genes in the giant panda （Ailuropoda mela- noleuca） and tested for selection effect and recombination event across exonic and intronic sequences to understand maintenance mechanism of polymorphism at Aime-MHC class I genes. In total, we isolated 21 MHC class I haplotypes （exon 2-intron 2-exon 3） from 46 captive giant pandas, of which eight were for Aime-C, seven for Aime-I and six for Aime-L; however, we only identified six unique sequences from these haplotypes. The subsequent maximum-likeli- hood and Chi-square analyses both detected evidence of recombination acting on the 21 haplotypes. These results indicate that the giant panda still retains a relatively high adaptive variation at Aime-MHC-I genes, and that the intronic segments have been homogenized along evolu- tionary time by recombination and subsequent genetic drift.We calculated nucleotide substitution rates of the antigen- binding regions （exons 2 and 3） and the noncoding intron 2, and found two pieces of evidence supporting the presence of balancing selection in the giant panda： an excess of nonsynonymous over synonymous substitutions at the antigen-binding sites, and an obviously higher synonymous substitutions in the exons than nucleotide substitutions in the intron. Thus, this study reveals that balancing selection and recombination together shape the diversity pattern at Aime- MHC-I loci of the giant panda.展开更多
文摘目的通过对1例新生儿期特殊面容、神经系统结构畸形患儿进行全外显子组序列检测分析,旨在为该患儿寻找潜在的致病原因。方法纳入1例在复旦大学附属儿科医院(我院)新生儿病房住院期间未能明确诊断的多发畸形患儿,主要临床表型为前额突出、腭弓高、耳位低、枕部较平,双侧脑室扩大、透明隔部分缺如、胼胝体发育异常,采用Sure Selct Human All Exon捕获试剂盒和Illumina Hi Seq2000测序平台,行全外显子组序列检测。数据分析采用复旦大学附属儿科医院转化中心所建立的高通量测序数据分析流程。采用Sanger测序进行验证。结果患儿全外显子组序列检测数据,共检测到79 064个变异,经过质量控制筛选、变异频率筛选、变异分类筛选,剩余645个变异。在进一步分析中,645个变异中有159个其所在基因在OMIM数据库及HGMD数据库与疾病相关。从3个已经报道的突变位点中锁定致病突变为FGFR2基因(NM_000141)c.C1040G,p.S347C。Sanger测序在家系内验证该位点为新发(de novo)突变。结论采用全外显子组序列检测,明确诊断FGFR2相关疾病1例。并且结合我院已经建立的高通量测序数据分析和临床诊断流程,为新生儿多发畸形寻找潜在的致病基因提供了快速、高效的方法。
文摘目的通过对1例智力发育迟缓患儿全外显子组测序(WES),旨在为该患儿寻找潜在的致病原因。方法纳入1例在复旦大学附属儿科医院(我院)住院期间诊断不明患儿,采用Sure Selct Human All Exon捕获试剂盒和Illumina Hi Seq2000测序平台,行WES检测;数据分析采用我院分子诊断中心建立的高通量测序数据分析流程;结果采用Sanger直接测序法进行验证。建立体外大鼠神经元原代培养和糖氧剥夺(OGD)模型,分为空白对照组(Sham+PBS亚组、Sham+POSTN亚组)、OGD组(OGD+PBS亚组和OGD+POSTN亚组),采用免疫荧光、LDH毒性检测、Brd U和TUNEL方法检测POSTN对2组及其亚组神经元生长增殖和凋亡的影响。结果患儿WES共检测到532 151个变异,筛选后检测到POSTN基因(NM_006475)外显子23:c.A2475T:p.G825G和外显子19:c.G2251A:p.E751K。Sanger测序显示p.G825G来自母亲,p.E751K来自父亲,符合复合杂合遗传模式。培养的大鼠原代神经元(MAP-2或Neu N阳性细胞)中有内源性POSTN的表达。LDH活性Sham+POSTN亚组与Sham+PBS亚组差异无统计学意义,OGD+POSTN亚组与OGD+PBS亚组差异有统计学意义。OGD+PBS亚组致神经元细胞缺氧2、6、12 d后比Sham+PBS亚组Brd U阳性的神经元数量增高,TUNEL阳性神经元凋亡也增高;OGD+POSTN亚组各时间点神经元增殖较OGD+PBS亚组均明显增高,但凋亡减少。结论 POSTN可以减少脑损伤后LDH对于神经元的细胞毒性,促进损伤后神经元的增殖,抑制凋亡。
基金supported by a special grant(SG1411)for the giant panda from the State Forestry Administration of China
文摘Major histocompatibility complex （MHC） is a family of highly polymorphic genes activating adaptive immunity in vertebrates. However, the underlying mecha- nism of MHC evolution is still not fully understood. In this study, we investigated genetic variation of three classical MHC class I genes in the giant panda （Ailuropoda mela- noleuca） and tested for selection effect and recombination event across exonic and intronic sequences to understand maintenance mechanism of polymorphism at Aime-MHC class I genes. In total, we isolated 21 MHC class I haplotypes （exon 2-intron 2-exon 3） from 46 captive giant pandas, of which eight were for Aime-C, seven for Aime-I and six for Aime-L; however, we only identified six unique sequences from these haplotypes. The subsequent maximum-likeli- hood and Chi-square analyses both detected evidence of recombination acting on the 21 haplotypes. These results indicate that the giant panda still retains a relatively high adaptive variation at Aime-MHC-I genes, and that the intronic segments have been homogenized along evolu- tionary time by recombination and subsequent genetic drift.We calculated nucleotide substitution rates of the antigen- binding regions （exons 2 and 3） and the noncoding intron 2, and found two pieces of evidence supporting the presence of balancing selection in the giant panda： an excess of nonsynonymous over synonymous substitutions at the antigen-binding sites, and an obviously higher synonymous substitutions in the exons than nucleotide substitutions in the intron. Thus, this study reveals that balancing selection and recombination together shape the diversity pattern at Aime- MHC-I loci of the giant panda.
