p表型个体的血清学特点和分子机制研究1例

目的分析1例p表型个体的血清学特征和分子机制。方法选取2021年5月在嘉兴市中心血站进行血型鉴定的1例p表型个体为研究对象。用血型血清学方法鉴定其ABO、RhD、P1PK血型和意外抗体。采用PCR-直接测序法(PCR-SBT)对编码P1和PK抗原的α1, 4-半乳糖基转移酶基因(A4GALT)的编码区进行测序分析。结果该个体的血型为A型、RhD阳性、P1PK系统为罕见的p表型, 血清中存在抗-PP1Pk。测序结果显示其A4GALT基因编码区存在c.343A>T纯合变异。结论 A4GALT基因c.343A>T纯合变异很可能导致了p表型个体。     

一个罕见葡萄糖转运体1缺陷综合征家系的遗传学分析

目的 对一个罕见葡萄糖转运体1缺陷综合征（GLUT1-DS）家系的致病基因变异位点进行检测并对其进行遗传学分析。方法 使用高通量全外显子组及PCR-Sanger检测家系成员的相关可疑突变位点,用I-TASSER生物信息学软件对含突变的蛋白质进行生物信息学分析,并采用PCR及毛细管电泳检测该核心家系成员中突变位点的嵌合比例。结果 先证者在2月龄时发生首次抽搐,且具有容貌异常、脑脊液葡萄糖含量较低和脑电图异常等临床特征。先证者及其母亲在SLC2A1基因（NM＿006516）检出杂合性无义突变c.988C>T(p.R330X),其母亲突变为新生突变（DNMs）,但未产生明显的临床表型;蛋白结构生物信息分析表明p.R330X无义突变所产生的截短蛋白使SLC2A1基因编码的GLUT1蛋白丧失部分功能;嵌合体分析显示先证者及其母亲的嵌合率相近。结论 该例GLUT1-DS患者的致病突变基因为SLC2A1基因c.988C>T无义突变,先证者母亲临床表型外显不全的原因可能是因为存在嵌合突变以外的分子病理机制。     

一例α1,4半乳糖基转移酶基因一碱基缺失导致p表型

目的　研究 α1,4半乳糖基转移酶基因决定 p表型的分子基础。　方法　在标准血清学鉴定红细胞表型的基础上 ,PCR扩增 p表型个体 α1,4半乳糖基转移酶基因的第 3外显子编码区序列 ,PCR产物经割胶纯化后直接进行测序分析。结果　p表型个体 α1,4半乳糖基转移酶基因第 3外显子第 30 0或30 1位核苷酸 G缺失 (TCGG→ TCG) ,导致阅读框架在第 10 1位氨基酸发生移码 ,在第 113位氨基酸处提前形成终止密码。先证者父母为杂合缺失携带者。　结论　发现了 α1,4半乳糖基转移酶基因第 3外显子第 30 0或 30 1位处 G缺失突变 ,该突变可能是 p表型的分子机理之一。     

一个遗传性凝血因子Ⅴ缺陷症家系表型与基因突变分析

目的 对一个由F5基因复合杂合突变导致的遗传性凝血因子Ⅴ（FⅤ）缺陷症家系进行表型和基因型分析,探讨其分子致病机制。方法 检测先证者及其家系成员（共3代7名成员）凝血酶原时间（PT）、活化部分凝血活酶时间（APTT）、纤维蛋白原（FⅠB）、凝血酶时间（TT）、血浆FⅤ活性（FⅤ:C）、FⅤ抗原（FⅤ:Ag）及其他相关凝血指标以明确诊断。通过自动校正凝血酶曲线法检测凝血酶生成量;用DNA直接测序法分析先证者F5基因的全部外显子、侧翼、5’和3’非翻译区及家系成员相应的突变位点区域;并用相应的生物信息学软件分析突变对蛋白功能的影响。结果先证者PT和APTT较正常对照明显延长,分别为17.9s/13.2s和46.9s/36s;FⅤ:C和FⅤ:Ag显著下降,分别为24%和28%;其父亲(Ⅰ1)、母亲(Ⅰ2)、妹妹(Ⅱ3)和儿子(Ⅲ1)FⅤ:C和FⅤ:Ag水平均有不同程度下降。基因分析显示,先证者第13号外显子存在c.2390＿2390delC杂合缺失突变（p.Pro798Leufs*13）以及第25号外显子存在c.6665A>G杂合错义突变（p.Asp2222Gly）;其父亲和妹妹为c....     

一例罕见β地中海贫血基因突变及其家系分析

 目的 鉴定1种罕见的β地中海贫血突变类型。方法 血液学分析采用血细胞分析仪及全自动快速电泳分析系统;α珠蛋白常规突变检测采用Gap-PCR;β珠蛋白常规突变检测采用反向点杂交法;样品的基因突变及基因型用β珠蛋白基因全长测序技术确定。 结果 先证者具有典型的β地中海贫血临床特点和血液学特性,HbF为 5.8%,其父母各项指标均正常。未发现先证者及其家庭成员有已知的α-/β-地中海贫血基因突变,测序发现先证者及其母亲均为CD2(CAT-CAC)杂合子,父亲为CAC纯合子;先证者有β珠蛋白exon1 CD27 (GCC-GAC)突变,编码的氨基酸由丙氨酸变为天冬氨酸,其未发现父母有CD27突变。结论 CD27(GCC-GAC)突变是罕见的β珠蛋白基因点突变,有助于指导人群筛查、遗传咨询和临床诊断。     

两个3型遗传性血管性血友病家系表型和基因型分析

目的 对2例遗传性血管性血友病家系进行临床表型诊断和基因型分析,探讨其分子发病机制.方法 分别采用出血时间、活化部分凝血活酶时间、瑞斯托霉素诱导的血小板聚集试验、血管性血友病因子(von Willebrand factor,vWF)瑞斯托霉素辅因子试验、vWF抗原、vWF活性、vWF胶原结合实验和多聚体分析对2例遗传性血管性血友病先证者和家系成员进行表型诊断.抽提先证者外周血基因组DNA,PCR法扩增VWF基因的52个外显子及侧翼序列,通过直接测序分析VWF基因突变.结果 2例先证者出血时间和活化部分凝血活酶时间均延长,血浆瑞斯托霉素诱导的血小板聚集、vWF瑞斯托霉素辅因子活性、vWF抗原、vWF活性、vWF胶原结合功能均显著降低,多聚体分析显示2例先证者多聚体均缺无,先证者A基因分析发现第17外显子的纯合插入突变g.82888_82889insCATG,导致740位蛋氨酸往后编码10个异常氨基酸后终止.先证者B基因分析发现第20外显子g.94865G＞A(Trp856X)和第28外显子g.110698_110699delinsG导致1481位甘氨酸往后编码42个异常氨基酸后终止的复合杂合突变.结论 g.82888_82889insCATG纯合插入突变和g.94865G＞ A(Trp856stop)与g.110698_110699delinsG的复合杂合突变分别导致了2例先证者的3型遗传性血管性血友病.     

中国人Wilson病ATP7B基因Thr935Met突变与临床表型的关系研究

目的　探讨Wilson病 (Wilsondisease,WD)ATP7B基因突变的高频位点之一Thr 935Met与临床表现的关系。方法　应用PCR技术扩增 90例WD患者和 30例正常人ATP7B基因第 12外显子 ,其PCR产物行限制性内切酶TaiI酶切分析。结果　正常人组酶切未见异常。 90例WD组第 12外显子有 10例Thr 935Met杂合突变 ,未见纯合突变 ,检出率为 11 1%。突变组的平均发病年龄为 19 10岁± 8 5 2岁 ,无突变组的平均发病年龄为 14 0 0岁± 7 4 2岁 ,差异有显著意义 (P =0 0 4 7)。结论　Thr 935Met突变组患者的发病年龄迟于未见该点突变组的患者 ,而性别、首发症状及铜生化水平与该点突变无明显相关性。     

一例罕见的新的TCIRG1基因杂合性突变引起的婴儿恶性石骨症

目的:研究1例婴儿恶性石骨症患者的致病基因及其突变。TCIRG1和CLCN7是婴儿恶性石骨症最常见的致病基因。前者被认为是纯合性致病基因,国外只有6例其杂合性改变也致本病的报导,而我国无杂合性突变导致本病的报道。方法:通过骨组织X线检查结合临床症状及体征确诊1例散发性婴儿恶性石骨症患者。提取患者及其父母的外周血基因组DNA,PCR扩增TCIRG1和CLCN7基因外显子及其剪切位点序列,对PCR产物直接测序。用TCRIG1基因附近的微卫星标记和SNP构建患者及其父母的单倍型。用染色体微阵列分析技术对患者及其母亲进行TCIRG1基因拷贝数目变异的检测。结果:患者TCIRG1基因第5号外显子内发现一个4个碱基的缺失突变c.449_452del AGAG(p.Gln149Glnfs16),该突变使得基因3’端编码的666个氨基酸被截断,失去了整个ATP酶V0复合结构域。患儿双亲TCIRG1和CLCN7基因的突变检测及单倍体构建证实该突变来源于患者父亲。染色体微阵列分析未发现患儿及其母亲携带有任何累及TCIRG1及CLCN7基因的拷贝数目变异。结论:本研究发现了1例TCIRG1基因新的杂合性突变所致的婴儿恶性石骨症。这是我国TCIRG1基因杂合性突变引起婴儿恶性石骨症的首例报道。这个发现可用于婴儿恶性石骨症的分子诊断。     

GJB2基因突变与临床表型关系的研究进展

GJB2基因是编码在耳蜗和表皮中表达的B类缝隙连接蛋白Connexin26（Cx26），该基因的突变被认为是语前遗传性非综合征性耳聋的分子遗传基础。GJB2基因与耳聋的关系密切，到目前为止，已发现GJB2基因有109种突变。这些突变所致的临床表型不一致，有些位点突变可导致耳聋，而有些只是基因的多态性；该基因的突变部位具有群体差异性。有关GJB2基因突变与临床表型的研究对遗传咨询和临床诊断具有重要的意义。     

六个成骨不全家系的临床表型分析及基因变异检测

目的分析6个成骨不全家系的临床表型并明确其致病变异,为遗传咨询及产前诊断提供依据。方法收集6个家系的临床资料以及外周血或引产组织样本,应用二代测序(next generation sequencing,NGS)技术对先证者的全部基因进行检测,用PCR反应扩增检出的变异位点,之后进行Sanger测序。在6个家系的所有成员以及100名健康对照中对检测到的变异位点进行验证。结果家系1的先证者及其女儿携带COL1A1基因c.1976G>C杂合变异,家系2~6的先证者分别携带COL1A2基因c.2224G>A、COL1A1基因c.2533G>A、COL1A2基因c.2845G>A、COL1A1基因c.2532_2540delCGGACCCGC以及COL1A2基因c.1847G>A杂合变异。先证者的双亲均未携带相应变异,在100名健康对照中均未检测到上述变异。结论6个成骨不全家系的致病原因可能均为COL1A1/2基因的变异。新发现的变异丰富了成骨不全症的表现型-基因型数据库,并为这些家系的遗传咨询及产前诊断提供了依据。     

一个Angelman综合征家系的临床表型及遗传学分析

目的:明确1个Angelman综合征(Angelman syndrome,AS)家系的遗传学病因,为家系的遗传咨询提供理论依据。方法:应用高通量测序技术进行单基因遗传智力障碍相关基因检测;应用拷贝数变异测序技术(copy number variation sequencing,CNV-seq)进行染色体非整倍体、100...     

The rare Lutheran blood group phenotype Lu(a–b–): a genetic study

The rare blood group phenotype lacking Lutheran antigens, Lu(a–b–), is known to have two genetic backgrounds. Tests on 250000 blood donors show the frequency of Lu(a–b–) to be approximately 1 in 3000. The families of 41 propositi show the dominant inhibitor of Lutheran antigens, In ( Lu ), to be the usual cause of the phenotype in South East England; there was no proven case of the recessive background, LuLu . Lod scores for In ( Lu ) and other blood group loci are presented; the only hint of linkage is between In ( Lu ) and Rh . The suppressing effect of In ( Lu ) on the expression of antigens of unrelated blood group systems, P₁, Au^a and i, is confirmed.     

一个遗传性癫痫伴热性惊厥附加症家系的临床表型及 SCN1A基因变异分析

目的:分析1个遗传性癫痫伴热性惊厥附加症(genetic epilepsy with febrile seizures plus,GEFS+)家系的临床表型及遗传特征进行分析。方法:收集先证者及其家系成员的临床资料并提取外周血DNA,对先证者采用高通量测序以确定潜在的变异位点,对其家系成员采用Sanger测序进行验证。...     

The estimation of phenotype distributions from pedigree data

A method is described to estimate from pedigree data the probability density function of a quantitative phenotype, given a specific genotype. Theoretically, pedigrees of any size or complexity that are segregating for conditions of known Mendelian inheritance can be used. Under certain circumstances, the estimates are unbiased by the method of ascertainment.     

类孟买表型 FUT1新等位基因的鉴定及家系研究

目的:报告1例类孟买血型并探讨其分子机制。方法:对1例ABO血型常规检测正反定型不相符的就诊者,采用吸收放散试验检测红细胞弱表达的ABH血型抗原,唾液酸实验检测唾液中血型物质。采用PCR产物直接测序法进行 ABO基因第6、7外显子和 FUT1和 FUT2基因序列分析。 结果...     

Developing a national collaborative study system for rare genetic diseases

There are thousands of rare genetic diseases and many genetic and nongenetic contributors to common genetic diseases. The evidence base that is currently available about the great majority of these conditions is limited to case studies and relatively small observational study sets derived from one or several institutions. Hence, the statistical power in any one study is usually quite limited. Further, in the absence of organized registries and data collection on particular patient groups, the information available is weak and the patient resources that are available are limited. It is only through organized and coordinated clinical investigation systems that a sufficient number of patients with these diseases can be accumulated to provide the statistical power needed to inform about clinical history of treated and untreated forms, provide the resources needed for clinical trials of new tests and treatments, provide a sufficiently powered evidence base for public health decision-making and other uses. The meeting in which these issues were raised resulted in a set of proposed principles and associated recommendations as to how best to achieve the vision of creating an extensive and comprehensive collaboration of professional and lay communities to enable translational research to improve clinical care and therapies for persons with rare genetic diseases.     

Creatine depletion in a new case with AGAT deficiency: clinical and genetic study in a large pedigree

Arginine:glycine amidinotransferase (AGAT, EC 2.1.4.1) deficiency is a recently recognized autosomal recessive inborn error of creatine biosynthesis, characterized by mental retardation and severe language impairment. We extensively investigated a third 5-year-old patient with AGAT deficiency, discovered in the pedigree of the same Italian family as the two index cases. At the age of 2 years he presented with psychomotor and language delay, and autistic-like behavior. Brain MRI was normal, but brain 1H-MRS disclosed brain creatine depletion, which almost completely normalized following creatine monohydrate supplementation. A remarkable clinical improvement paralleled the restoration of brain creatine concentration. AGAT and GAMT (guanidinoacetate:methyltransferase) genes were analyzed in the proband and in 26 relatives, including the two cousins with AGAT deficiency. Sequencing of the proband's AGAT gene disclosed the same homozygous mutation at nt position 9093 converting a tryptophan (TGG) to a stop codon (TAG) at residue 149 (W149X), as already described in the two previously reported cases. The proband's parents and 10 additional subjects of the pedigree were carriers for this mutation. AGAT deficiency was further confirmed by undetectable AGAT activity in the patient's lymphoblasts. Mutation analysis of the GAMT gene revealed a sequence variation in exon 6 (T209M), not in the proband, but in 15 additional subjects from the pedigree. The silent nature of this sequence variation is supported by its homozygosity in one AGAT deficient cousin and in one asymptomatic adult, both with normal GAMT activity.     

The relevance of pre-amniocentesis pedigree analysis and genetic counseling

Prenatal diagnosis by amniocentesis in the second trimester of pregnancy has become widely accepted, and the demand for the procedure is increasing exponentially. It is important to reevaluate critically the time and effort spent obtaining a detailed pedigree analysis and family history prior to amniocentesis. Two hundred unselected consecutive cases of women undergoing amniocentesis because of advanced maternal age were studied. One woman had a brother with Duchenne muscular dystrophy and she was found by CPK testing to be a carrier. The prenatal diagnosis revealed that she was carrying a male fetus with a 50% chance of being affected, and the couple decided to have a termination of the pregnancy. A variety of other familial disorders and teratogenic exposures which were found in the studied 200 families were further explored through genetic counseling. Additional reasons for the individual pre-amniocentesis counseling include reducing anxiety and responding to specific psychosocial aspects of the prenatal diagnosis procedure in particular families.     

一个X-连锁肾上腺脑白质营养不良家系的临床特点及基因变异分析

目的:探讨一个X-连锁肾上腺脑白质营养不良家系的临床特点及 ABCD1基因的变异方式。 方法:分析X-连锁肾上腺脑白质营养不良家系的临床资料,采用PCR方法对家系中的先证者、父母以及100名健康对照者的 ABCD1基因进行DNA测序。 结果:患者主要以脑干、小脑损害为突出表现...     

硫胺素焦磷酸激酶缺乏症一家系的临床特点及基因变异分析

目的:探讨一个硫胺素焦磷酸激酶缺乏症(thiamine pyrophosphokinase deficiency,TPKD)家系的临床和基因变异特点。方法:分析TPKD家系的临床表现、影像学特点,对家系成员行全外显子测序。结果:先证者,女,2岁8个月出现发作性共济失调伴肌张力障碍,此后反复发作共济失调7～8次。11岁时...