Hemophilia B: diagnostic value of RFLP analysis in 19 of the 20 known Finnish families

With the aim of determining the usefulness of RFLP analysis in carrier detection and prenatal diagnosis, we studied all available members including 40 patients, 30 obligate carriers and 39 women at risk belonging to 19 out of a total of 20 Finnish hemophilia B families. The allele frequencies of the three intragenic polymorphisms studied (TaqI, XmnI and DdeI) did not differ significantly from those reported in other Caucasian populations. A considerable degree of linkage disequilibrium between the three polymorphisms was observed. Carriership evaluated in 39 females at risk led to exclusion in 14 while carriership was established in 5. The proportion of women who by pedigree analysis had a carriership risk between 10% and 90% could be reduced from 97% to 51% by RFLP analysis. Prenatal diagnosis using an intragenic polymorphism could potentially be offered to 69% of hemophilia carriers. DNAs from 19 unrelated patients were screened for mutations using a full-length cDNA probe, but no abnormal hybridization patterns were observed. Our results indicate that RFLP segregation analysis provides a useful method of carrier detection and prenatal diagnosis in hemophilia B.     

Marfan syndrome: exclusion of genetic linkage to the COL1A2 gene

Marfan Syndrome is a genetic disorder of the connective tissue. Individuals from one large family with this disorder were genotyped for COL1A2 gene associated RFLPs. Our results demonstrated that the COL1A2 gene, encoding the proa2(I) collagen chain, segregated independently of the phenotype and it is therefore excluded as the mutant locus in this family.     

RFLP analysis for diagnosis of haemophilia A in the German Democratic Republic

The frequencies of Bcl I, Hind III and Xba I intragenic polymorphic sites in the population of the GDR were found to be 0.68, 0.38 and 0.48, respectively. No differences in composition and frequencies were detectable at DXS 52 locus in comparison with other Caucasian populations. A strong linkage disequilibrium between the intragenic Bcl I and Hind III sites could be confirmed. The observed heterozygosity for the flanking marker DXS 52 in combination with intragenic Bcl I and Xba I polymorphisms was 0.97. Using these three RFLPs, 122 females at risk in 41 independent haemophilia A families were investigated; 86 of them could be identified and 27 excluded as carriers; 9 females could not be classified. So far, four prenatal diagnoses in the first trimester of gestation have been performed by RFLP analysis.     

Analysis of fragile X-mental retardation families using flanking polymorphic DNA probes

Fragile-X mental retardation (FRAX-MR) is one of the more common X-linked disorders affecting 1 in 1,500 newborn males. This disease is characterized by the expression of fragile site in the region q27.3 of the X-chromosome of affected boys when their lymphocytes are cultured in folate deficient medium. In most patients there is macroorchidism postpubertally. The clinical diagnosis of carrier females based on the expression of fragile site in Xq27.3 is usually difficult and sometimes impossible. About half of the carrier females escape diagnosis by this method. Furthermore, prenatal diagnosis is not always feasible. Using Restriction Fragment Length Polymorphism (RFLP) and cloned DNA segments from the region Xq27-Xqter as probes, we have investigated Swedish families with FRAX-MR in three generations. Interesting observations, previously unreported to our knowledge, have been made in some patients and carrier mothers, using one of the probes which is localized to the distal end of Xq. The significance of these findings and the linkage of the disease locus to the different probes used in this study is presented.     

Multipoint linkage mapping of the Xq25-q26 region in a family affected by the X-linked lymphoproliferative syndrome

We have performed, in a large Swiss family, a study of linkage between various DNA markers in the Xq24-27 region and the locus for the X-linked lymphoproliferative syndrome (XLP). Our results indicated that the marker DXS37 in Xq25-q26 is genetically linked to the XLP syndrome. The multipoint linkage analysis showed that the disease locus is distal to DXS11, but proximal to the hypoxanthine phosphoribosyl-transferase gene (HPRT).     

血友病甲基因分析技术的改进及其在产前诊断中的应用

目的对血友病甲基因分析技术进行改进并应用于携带者检查和产前诊断。方法长距离聚合酶链反应方法直接检测凝血因子Ⅷ第22内含子倒位,对非倒位家系用FⅧ基因内限制酶切位点XbaⅠ、HindⅢ、二核苷酸重复序列多态性位点STR13和STR22,以及基因外可变数目串联重复序列DXS52(St14)位点进行基因连锁分析。结果52个家系共检出71位携带者。21个家系为第22内含子倒位,28个家系经连锁分析得到明确诊断,3个家系无法诊断,可诊断家系占94.2%。为18个家系做胎儿产前诊断,其中10例诊断为血友病甲胎儿;诊断7例正常男胎和1例携带者女胎,随访1年发育正常。结论应用长距离聚合酶链反应和多位点基因连锁分析技术可以快速有效地进行血友病甲携带者检查和产前诊断。     

Use of the RB1 cDNA as a diagnostic probe in retinoblastoma families

Use of an intragenic BamHI restriction fragment length polymorphism within the 5' end of the retinoblastoma gene (RB1) provided improved genetic counselling for five familial and ten non-familial retinoblastoma patients and their relatives. All other polymorphic probes within RB1 were uninformative in three families, and accuracy of diagnosis was improved by use of this polymorphism in two families. In 10/14 informative constitutional DNA-RB tumor DNA pairs, a reduction to homozygosity allowed identification of the RB1 allele at risk to carry a germline RB1 mutation.     

Genome-wide linkage analysis is a powerful prenatal diagnostic tool in families with unknown genetic defects

Genome-wide linkage analysis is an established tool to map inherited diseases. To our knowledge it has not been used in prenatal diagnostics of any genetic disorder. We present a family with a severe recessive mental retardation syndrome, where the mother wished pregnancy termination to avoid delivering another affected child. By genome-wide scanning using the Affymetrix (Santa Clara, CA, USA) 10k chip we were able to establish the disease haplotype. Without knowing the exact genetic defect, we excluded the condition in the fetus. The woman finally gave birth to a healthy baby. We suggest that genome-wide linkage analysis – based on either SNP mapping or full-genome sequencing – is a very useful tool in prenatal diagnostics of diseases.     

PCR-构象敏感凝胶电泳技术在血友病A基因分型诊断及携带者检测中的应用

所有的HA家系(含散发)进行直接基因诊断,理论上可筛查新突变并明确其突变类型.该法简便、快速、成本低,在HA直接基因诊断及携带者筛查中优势独特,应具重要应用价值.     

Genetic heterogeneity in X-linked agammaglobulinemia complicates carrier detection and prenatal diagnosis

X-linked agammaglobulinemia (XLA) is a severe antibody deficiency disease reflecting an arrest of B lymphocyte differentiation at the level of precursor B cells. The disease is inherited in an X-linked recessive mode. In a single eight-generation pedigree the XLA gene was mapped to the Xq21.3-Xq22 area of the X chromosome. The data establish close linkage of the XLA locus to the DXS17 restriction fragment length polymorphic (RFLP) marker locus (the lod score exceeding 6 at phi = 0). A series of RFLP markers around the DXS17 locus provided an RFLP haplotype of use in genetic counselling within this pedigree. In one other pedigree a phenotypically identical disease was inherited but was accompanied by a high frequency of recombination with the DXS17 locus, which made localisation of the gene at the DXS17 locus highly unlikely (lod score less than -3). This genetic heterogeneity complicates genetic counselling within particular pedigrees, especially when the localization of the XLA gene involved in those pedigrees has not been established.     

DNA polymorphisms and deletion analysis of the Duchenne–Becker muscular dystrophy gene in the chinese

In our investigation of Duchenne muscular dystrophy (DMD)–Becker muscular dystrophy (BMD)gene in the Chinese, the analysis of relevant restriction fragment length polymorphisms (RFLPs) was first made in 30 normal female volunteers to determine their allele and genotype frequencies, and then in 29 DMD–BMD families for informativeness of different combinations of RFLPs in making carrier detection and prenatal diagnosis. We further screened the mutant gene, first with four 5' end intronic, genomic probes (pERT87-1, pERT87-8, pERT87-15, and XJ1.1) which did not show any deletions, and then with all dystrophin cDNA probes which disclosed 13 partial gene deletions out of 29 patients studied (45%). The deletions were non-randomly distributed, clustering primarily near the central region of the gene. Fifty percent of the deletions involved single exon-containing HindIII restriction fragments, and again most were located near the center of the gene, emphasizing the importance of this area. Some exceptions were found against the previous suggestion that intactness of translational open reading frame resulted in a BMD phenotype. Neither the location of the break-points nor the length of the deletions was useful in predicting a certain phenotype. One of our patients had an intriguing pattern of partial gene deletion that lost part of the gene at the 3' end. Carrier determination was attempted by use of dosage analyses or identification of junction fragments which greatly improved accuracy and reliability.     

血管紧张素原基因单倍型与原发性高血压的关联研究

目的 研究中国人血管紧张素原（angiotensinogen,AGT)基因多态性及单倍型与原发性高血压的关系。方法 在335例高血压患者与196名血压正常人中采用PCR－限制性酶切片段长度多态性法检测血管紧张素原基因的多态性，同时用最大期望值方法进行两位点连锁不平衡和三位点的单倍型分析。结果 在M235T和A－20C，M235T和A－6G，A－20C和A－6G位点观察到了连锁不平衡（P＜10^-4)。病例－对照检验显示T235等位基因频率在高血压组中高于对照组，但所有单倍型频率分布在高血压组和正常对照组间差异无显著性。结论 受检人群中AGT基因各单倍型与高血压未见关联，但AGT基因T235位点可能是一种重要的高血压风险因子。     

Analysis of restriction fragment length polymorphism of cytochrome P450 3A43 gene and evaluation of the incidence of <Emphasis Type="Italic">CYP3A43*1B</Emphasis> allele in europeoid residents of West Siberia

Analysis of restriction fragment length polymorphism is proposed for detecting CYP3A43 gene c1047>T mutation. The incidence of this mutation (CYP3A43*1B allele) was evaluated in 102 europeoid residents of West Siberia and it was found to be 12.25%. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 140, No. 12, pp. 681–683, December, 2005     

Lack of association between TaqI A1 allele of dopamine D2 receptor gene and alcohol-use disorders in Atayal natives of Taiwan

Association studies between the A1 allele of the dopamine D2 receptor (DRD2) gene TaqI A polymorphism and alcoholism remain controversial. A recent study from Japan demonstrated that the A1 allele is associated with severe alcoholism in the Japanese population. We were interested in knowing if this association also exists in the Atayals of Taiwan, who were found to have a higher prevalence of alcohol-use disorders than the Han Chinese in Taiwan. Genotype and allele frequencies were determined in alcohol-abusing, alcohol-dependent, and nonalcoholic control Atayal natives in Taiwan. A1 allele frequencies in alcohol-dependent, alcohol-abusing, and normal control Atayals were 0.39, 0.42, and 0.39, respectively. No difference in A1 allele frequency was found among these three groups. Our data do not support the hypothesis that the A1 allele of the TaqI A polymorphism of the DRD2 gene increases susceptibility to alcohol-use disorders in the Atayals of Taiwan. © 1996 Wiley-Liss, Inc.     

First trimester prenatal diagnosis of haemophilia a using factor VIII gene probe

Accurate first-trimester prenatal diagnosis was achieved in a Japanese haemophilia A family by the use of a restriction fragment length polymorphism (RFLP) located within the F.VIII gene. Since the pregnant woman's heterozygosity forBclI polymorphism in F.VIII/intron 18 (F8A) probe was informative, chorionic villus sampling (CVS) was performed at 9 weeks of gestation. Restriction analysis showed that the fetus was heterozygous for theBclI site and had received a normal paternal X chromosome (0.9 kb) and a normal maternal X (1.2 kb). Therefore, we concluded that the fetus was a non-carrier female. Pregnancy went to term and woman gave birth to an apparently healthy female. At one week after birth a coagulation study confirmed that the newborn infant is not a carrier. The first-trimester prenatal diagnosis of haemophilia A is possible by CVS due to a RFLP in the F.VIII gene.     

Hemophilia A and B — two years ex-perience of genetic counselling and prenatal diagnosis

Haemophilia A . Thirty-one pregnant women, obligate or probable carriers of haemophilia A, requested prenatal diagnosis if sex determination showed the foetus to be a male. In 11 of the 31 cases the foetuses were females; in two, the genetic variant of the disease rendered prenatal diagnosis impossible; and in two, the mother aborted spontaneously. From the remaining 16 male foetuses, blood samples were obtained in uteri in the 17th to 20th week of gestation. Examination of the samples showed that 11 of the foetuses were unaffected and five affected. Haemophilia B . Three carriers of haemophilia B had male foetuses. Examination of foetal blood obtained in utero showed that these three foetuses were affected. Confirmation . All women with an affected foetus requested termination of pregnancy. In one of thecdses of abortion, no blood was obtained for confirmative examination. In the remaining cases, the prenatal prediction was confirmed in the abortus or in the child after birth; three women are still pregnant.     

β-Thalassaemia mutations and their linkage to β-haplotypes in Tamil Nadu in Southern India

A study for screening of beta-thalassaemia mutations by the Amplification Refractory Mutation System (ARMS) and haplotyping by Polymerase Chain Reaction (PCR) was undertaken because there was a paucity of data in Tamil Nadu in Southern India and to initiate a comprehensive prenatal diagnosis programme. A total of 294 alleles were analysed to study the nature of the mutations, of which 146 were beta-thalassaemia alleles. Only four types of beta-thalassaemia mutations were recorded. Of these, 128 alleles were of the variant IVS-1 nt 5 (G-->C). Thirteen had the mutation codon 41/42 (del TCTT), four had the mutation codon 8/9 (insert G) and one had the 619 bp deletion at the 3' end of the gene. The most common mutation, IVS-1 nt 5 (G-->C), was strongly associated with a single haplotype although the association was not absolute. The population of Tamil Nadu in Southern India seems to be ideal for initiating a prenatal diagnosis programme based on direct detection of mutation by ARMS coupled with RFLP linkage analysis.     

单胺氧化酶A基因EcoRⅤ多态与帕金森病的关联分析

目的　研究单胺氧化酶 A(monoamine oxidase A,MAO- A)基因 Eco R 多态 (C/ T)位点在中国人群中的分布 ,并探讨其与帕金森病 (Parkinson's disease ,PD)发病风险的关系。方法　采用聚合酶链反应 -限制性片段长度多态性分析法 ,在 110例 PD患者和 182名正常人中分析了 MAO- A基因 Eco R (C/ T)多态的分布 ,并对该多态与 PD进行关联分析。结果　 (1) MAO- A基因 Eco R 多态与 PD间不存在明显关联 (χ2 =0 .0 91,P=76 .3) ;(2 ) MAO- A基因 Eco R 多态在中国汉族人群和北美高加索人群中的分布差异有显著性 (χ2 =30 .0 3,P=4 .18)。结论　 MAO- A Eco R 多态可能与中国人 PD的发病风险无关     

一个新的COL4A5基因的剪接位点突变引起Alport综合征

目的 对一个Alport综合征家系进行研究,期望找到导致该家系发病的遗传基础.方法 对家系成员采样并提取DNA,对家系中的先证者和1名正常对照进行COL4A5基因全部编码区域的突变检测,限制性片段长度多态件分析技术对家系中所有成员和200名正常对照进行验证.结果 在该Alport综合征家系中发现一个新的COL4A5基因的剪接位点突变c.1517-1G＞T,而在家系的未患病成员,以及对照人群中未能检测到该突变.结论 发现了一个新的COL4A5基因的剪接位点改变c.1517-IG＞T,该突变可导致Alport综合征,该发现丰富了引起Alport综合征的COL4A5基因的突变谱.