中国男性雄激素受体基因(CAG)n重复多态性的初步研究

目的 研究正常中国男性雄激素受体（androgen receptor,AR)基因（CAG）n重复序列的多态性。方法 应用DNA测序基础上的[α-^32P]dCTP掺入不对称PCR-变性聚丙烯酰胺凝胶电泳（Polyacrylamide gel electrophoresis,PAGE)方法,对107名正常男性AR基因的（CAG）n重复数进行测定。结果 AR基因（CAG）n序列在正常男性人群中呈现重复多态性,其重复范围为11-29,集中于20-24,以22最多。结论 AR基因（CAG）n序列在正常男性人群中呈现重复多态性,为今后进一步研究其病理学及遗传学意义打下基础。     

Bias in the genomic distribution of CAG and CTG trinucleotide repeats

We investigated the hypothesis that the trinucleotide repeat CAG is disproportionately located in exons in genomic DNA by analyzing unbiased genomic sequences with the Gene Recognition and Analysis Internet Link program ( http//avalon.epm.ornl.gov/ ). Forty percent of CAG/CTG repeats were predicted to lie within exons. This is significantly greater than would be expected by chance, and is also greater than we have observed for ATT/AAT repeats. Therefore, our data support the hypothesis. Furthermore, the data support the utility of a recently reported CAG/CTG PCR genomic screening set for identifying pathogenic expanded CAG/CTG repeats. Am. J. Med. Genet. 74:62–64, 1997. © 1997 Wiley-Liss, Inc.     

Correlation between the incidence of myotonic dystrophy in different groups in Israel and the number of CTG trinucleotide repeats in the myotonin gene

Myotonic dystrophy (DM) is associated with an increased number of CTG repeats in the 3′ untranslated region of the myotonin gene. Because DM has been observed less frequently in Ashkenazic Jews and non-Jews than in North African and Yemenite Jews in Israel, a study of the CTG repeat polymorphism was undertaken in these four groups. Alleles from 126 unrelated healthy North African Jews, 103 Yemenite Jews, 103 Ashkenazic Jews, and 106 Israeli Moslem Arabs were studied by PCR analysis of the trinucleotide repeat in the DM gene, and the size distribution of the CTG repeat was determined. The alleles ranged in length from 5–28 repeats in the Yemenite Jews, 5–26 in Muslim Arabs and North African Jews, and 5–23 in the Ashkenazic Jews. North African and Yemenite Jews were found to have significantly more large repeats in the normal range than Ashkenazic Jews and Muslim Arabs (for over 18 repeats: 9.1% and 13%, respectively, compared to 2.4% and 3.3%, respectively; P < 0.0001). It is suggested that the more frequent occurrence of large CTG repeats in the normal range may represent a greater predisposition to DM. Am. J. Med. Genet. 71:156–159, 1997. © 1997 Wiley-Liss, Inc.     

Alu repeats in the human factor IX gene: The rate of polymorphism is not substantially elevated

Previous data suggested an elevated rate of polymorphism in Alu sequences. Direct genomic sequencing was performed on five Alu repeats in the factor IX gene in at least 20 unrelated males of European and Asian descent (40 kb total). In these Alu sequences, the estimated rate of polymorphism in Caucasians (H_E = 7.1 × 10^?4; H_N = 4.5 × 10^?4) is similar to previously examined nonrepetitive sequences in the factor IX gene, and about twofold lower than previous estimates of the average rate of polymorphism for the X-chromosome which utilized random genomic clones to detect RFLPs. The aggregate data on the rate of polymorphism in Alu sequences suggest that mutations due to gene conversions at these sites are infrequent. © 1993 Wiley-Liss, Inc.     

Independent origin of single and double mutations in the human glucose 6-phosphate dehydrogenase gene

The vast majority of both polymorphic and sporadic G6PD variants are due to single missense mutations. In the four polymorphic variants that have two point mutations, one of the mutations is always 376 A→G (126 Asn→Asp), which on its own gives rise to the nondeficient polymorphic variant, G6PD A. In a study of G6PD deficient patients who presented with clinical favism in Spain, we have found a new polymorphic variant that we have called G6PD Malaga, whose only abnormality is a 542 A→T (181 Asp→Val) mutation. This is the same mutation as previously found in association with the mutation of G6PD A in the double mutant, G6PD Santamaria. G6PD Malaga is associated with enzyme deficiency (class III), and the enzymic properties of G6PD Malaga and G6PD Santamaria are quite similar, indicating that in this case the effects of the two mutations are additive rather than synergistic. G6PD Santamaria might have been produced by recombination between G6PD A and G6PD Malaga; however haplotype analysis, including the use of a new silent polymorphism, suggests that the same 542 A→T mutation has taken place independently in a G6PD B gene to give G6PD Malaga and in a G6PD A gene to give G6PD Santamaria. These findings help to outline the relationship and evolution of mutations in the human G6PD locus. © 1996 Wiley-Liss, Inc.     

汉族人群载脂蛋白CII基因二核苷酸串联重复序列(TG)n(AG)m及(AG)m多态性

采用套式聚合酶链反应结合变性聚丙烯酰胺凝胶电泳和银染技术，并构建载脂蛋白CII（ApoCII)基因二核苷酸串联重复序列（TG）n(AG)m及（AG）m序列等位基因梯阶标准；检测正常汉族人群基因型和等位基因频率分布，检出36种（TG）n(AG)m序列基因型、12种等位基因。等位基因为17、18、26－35，其频率分别为0．061、0．011、0．002、0．002、0．054、0．255、0．372、0．084、0．026、0．039、0．052、0．041。检出7种（AG）m序列基因型、4种等位基因。等位基因为6、7、8、9，其频率分别为0．002、0．152、0．812、0．034。与欧洲白种人比较，ApoCII基因二核苷酸串联重复序列（TG）n(AG)m及（AG）m序列等位基因频率分布均具有明显的种族差异性（P＜0．01，P＜0．01）。     

脆性X综合征FMR1基因CGG重复序列与甲基化的检测

目的建立一种快速、可靠的脆性X综合征的群体筛查方法。方法应用热启动PCR和甲基化特异性PCR（MS-PCR）方法对62例智力低下儿童、12例父母外周血液以及5例高危胎儿的脐带血中FMR1基因CGG重复序列与甲基化状态进行检测。结果采用热启动PCR方法检测79例标本,77例标本的CGG重复数在21～40之间,与正常对照组无明显差异;2例标本未扩增出明显条带。采用MS-PCR方法检测出2例FMR1基因甲基化但CGG重复数在正常范围的患者。结论应用热启动PCR结合MS-PCR方法检测FMR1基因CGG重复数和甲基化,能提高诊断效率,可作为筛查脆性X综合征的首选方法。     

Nine mutations in the cystic fibrosis (CF) gene account for 80% of the CF chromosomes in French patients

Thirteen mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene have been screened in a French sample of 185 cystic fibrosis (CF) patients, together with their respective associated RFLP haplotypes at the linked D7S23 locus (XV2C and KM19 markers). The respective frequencies of the mutations showed that 9 of them account for 80% of the CF chromosomes. Implications for prenatal diagnosis and heterozygote detection are defined and discussed. The well-known great excess of RFLP B marker within CF chromosomes is partially explained by two already characterized mutations highly associated with haplotype B: delta F508 and G542X. Similarly, the excess of haplotype D within CF chromosomes is partially explained by the association between delta I507 and this haplotype. These results may suggest the existence of two still untested or uncharacterized mutations, whose frequencies could be near 1%, one which would be associated with haplotype B and a second which would be associated with haplotype D. The possible cause of the specific association between most of the main different CF mutations and the RFLP haplotype B is discussed.     

Analysis of the (CAG)n repeat causing Huntington's disease in a Mexican population

We investigated the allele distribution of the polymorphic (CAG)n repeat in the IT15 gene in 96 normal subjects from the Mexican population and 83 unrelated patients with Huntington's disease. Our results show that the size distributions of normal and affected alleles do not overlap. Normal alleles range from 13 to 32 triplets, with 18 being the most frequent allele, while HD alleles contain 37 to 76 repeats with 42 being the most frequent. One allele in the range of intermediate alleles was found (32 repeats) in a normal subject. The juvenile onset cases in this study are associated with an expansion greater than 49 repeats. In the available parent-offspring pairs, paternal alleles show instability with an expansion of 28 repeats in one case.     

Cyclin E gene (CCNE) amplification and hCDC4 mutations in endometrial carcinoma

Cyclin E overexpression occurs in a subset of endometrial carcinomas (ECs), but the molecular mechanisms underlying this alteration remain to be established. The present study has analysed amplification of the cyclin E gene (CCNE) and mutation in hCDC4, the gene coding for the F-box protein, which tags phosphorylated cyclin E for proteosomal degradation, to ascertain whether these alterations might be responsible for cyclin E overexpression in ECs. Cyclin E and p53 expression was studied by immunohistochemistry in eight atypical endometrial hyperplasias (AEHs), 51 endometrioid endometrial carcinomas (EECs), and 22 non-endometrioid endometrial carcinomas (NEECs). CCNE amplification was analysed by fluorescence in situ hybridization (FISH). Mutations in exons 2-11 of the hCDC4 gene were screened by PCR-SSCP-sequencing. Finally, the polymorphic marker D4S1610 was used to assess loss of heterozygosity (LOH) in the hCDC4 gene. Cyclin E overexpression was found in 26/81 (32%) cases and was associated with the histological type of the lesion, since it was not found in any AEHs but was present in 27% of EECs and 54.5% of NEECs (p=0.035). Cyclin E overexpression was associated with histological grade (p=0.011) and p53 immunostaining in EECs (p=0.033). CCNE amplification was found in 6 of 37 (16%) ECs examined. There was a significant association between CCNE amplification and the histological type of the lesion, since five (83%) of the six cases with amplification were NEECs (p=0.008). One EEC harboured an hCDC4 mutation: a CGA to CAA (Arg/Gln) change at codon 479. In addition, D4S1610 LOH was found in 7 of 23 (30%) informative cases analysed, but no correlation with cyclin E overexpression was found. However, the tumour with hCDC4 mutation also showed LOH. This is the first study demonstrating that cyclin E overexpression is associated with gene amplification in ECs, these alterations being more frequent in NEECs. Although hCDC4 exhibits a low mutation frequency in ECs overexpressing cyclin E, it seems to function as a tumour suppressor gene that is involved in endometrial carcinogenesis.     

宁夏回、汉族群体雄激素受体基因(CAG)n、(GGN)n重复多态性研究

目的 研究雄激素受体(androgen receptor,AR)基因(CAG)n、(GGN)n重复序列多态性在宁夏回、汉族群体中的分布特征.方法 用ABI 3730XL测序仪测定AR基因的(CAG)n及(GGN)n重复数目并对结果进行分析.结果 CAG等位基因在宁夏回、汉族群体中的分布差异无统计学意义(P＞0.05).GGN等位基因在两个群体中的分布差异有统计学意义(P＜0.01),汉族女性GGN重复数为23的等位基因频率(48.4％)显著低于回族女性(64.7％,P=0.01).结论 AR基因GGN等位基因在宁夏回、汉族群体中的分布差异有统计学意义(P＜0.01).     

Novel germline mutations in the APC gene of Cypriot patients with familial and sporadic adenomatous polyposis

Familial adenomatous polyposis (FAP) is one of the two commonest familial syndromes that predispose to colorectal cancer. FAP is caused by mutations in the adenomatous polyposis coli (APC) tumour suppressor gene that has a high penetrance. The disease is characterized by the occurrence of hundreds to thousands of colorectal polyps, which if left untreated give rise to colorectal cancer. In Cyprus, there are no molecular data available as yet on families with FAP. This work presents the results of APC analysis in our population for the first time. The APC gene was analyzed in 33 DNA samples from 20 individuals belonging to four FAP families and 13 patients with sporadic polyposis. We identified three truncating mutations, four missense mutations and 11 polymorphisms. It is of interest that two of the three truncating mutations, 2307delA and Q1242X, are novel, which supports the existence of a unique genetic pool in the Cypriot population. This ethnic molecular study in addition to highlighting population heterogeneity also contributes to phenotype-genotype associations that are essential for the clinical management of FAP families in Cyprus.     

Novel and recurrent mutations in the AIRE gene of autoimmune polyendocrinopathy syndrome type 1 (APS1) patients

Autoimmune polyendocrinopathy syndrome type 1 (APS1) is characterized by the presence of at least two out of three clinical features, which include Addison's disease, hypoparathyroidism, and chronic mucocutaneous candidiasis. This disorder is caused by mutations in the AIRE (autoimmune regulator) gene. While several AIRE mutations have been described in APS1 patients of various ethnic origins, the genetic cause of APS1 in Arab patients requires further investigation. This study describes seven Arab families, in which 18 patients had APS1. In addition to the cardinal features of APS1, some patients exhibited alopecia, diabetes mellitus, nephrocalcinosis and other phenotypes associated with APS1. DNA sequencing of the AIRE gene of patients from this study identified four novel and one recurrent mutation. These mutations likely result in loss of AIRE function in the patients. In addition, it was noted that the non-pathogenic c.834C> G mutation (rs1800520, encoding for p.Ser278Arg) occurs with high incidence in the AIRE gene of Arab individuals. Furthermore, this investigation demonstrates inflammation of the hair follicles in APS1 patients with alopecia universalis. We conclude that Arab APS1 patients carry novel and recurrent mutations in the AIRE gene.     

Transcervical cells and the prenatal diagnosis of haemoglobin (Hb) mutations

Prenatal diagnoses of haemoglobin (Hb) mutations were performed using transcervical cells, retrieved by aspiration from the endocervical canal of ten selected pregnant women at about 10 weeks of gestation, prior to chorionic villus sampling (CVS). Both parents were carriers of haemoglobinopathies (thalassaemia or HbS). Clumps of fetal cells were isolated by micromanipulation under an inverted microscope and aliquots of the extracted DNA tested separately for the presence of paternally derived chromosome markers and Hb mutations by quantitative fluorescent polymerase chain reaction (PCR). The correct prenatal diagnosis of Hb diseases, using selected single clumps of trophoblastic cellular elements free of maternal contaminating cells, was achieved in six out of ten cases.     

DNA deletions in the low density lipoprotein (LDL) receptor gene in Danish families with familial hypercholesterolemia

DNA samples from 25 unrelated Danish patients with familial hypercholesterolemia (FH) were screened by Southern blot hybridization to detect gross alterations in the low density lipoprotein (LDL) receptor gene. Three FH-patients were found to have a deletion. Two of these delete part of the cysteine rich domain, which comprises the ligand binding region of the LDL-receptor. The third deletion encompasses coding regions for the cytoplasmic part of the receptor. As two of these deletions could be equivalent to previously described LDL-receptor gene alterations, these data seem to support a notion of recombination hot spots which involve Alu-sequences.