Localization of cystic fibrosis locus to human chromosome 7cen-q22

Cystic fibrosis (CF) is the most common genetic disease in Caucasian populations, with an incidence of 1 in 2,000 live births in the United Kingdom, and a carrier frequency of approximately 1 in 20. The biochemical basis of the disease is not known, although membrane transport phenomena associated with CF have been described recently. Consanguinity studies have shown that the inheritance of CF is consistent with it being a recessive defect caused by a mutation at a single autosomal locus. Eiberg et al. have reported a genetic linkage between the CF locus and a polymorphic locus controlling activity of the serum aryl esterase paraoxonase (PON). The chromosomal location of PON, however, is not known. Linkage to a DNA probe, DOCR1-917, was also recently found at a genetic distance of approximately 15 centimorgans (L.-C. Tsui and H. Donnis-Keller, personal communication), but no chromosomal localization was given. Here we report tight linkage between the CF locus and an anonymous DNA probe, pJ3.11, which has been assigned to chromosome 7cen-q22.     

A closely linked genetic marker for cystic fibrosis

Cystic fibrosis is a recessive genetic disorder, characterized clinically by chronic obstructive lung disease, pancreatic insufficiency and elevated sweat electrolytes; affected individuals rarely live past their early twenties. Cystic fibrosis is also one of the most common genetic diseases in the northern European population. The frequency of carriers of mutant alleles in some populations is estimated to be as high as 1 in 20, carrying a concomitant burden of about one affected child in 1,500 births. Because little is known of the essential biochemical defect caused by the mutant gene, a genetic linkage approach based on arbitrary genetic markers and family studies is indicated to determine the chromosomal location of the cystic fibrosis (CF) gene. We have now obtained evidence for tight linkage between the CF locus and a DNA sequence polymorphism at the met oncogene locus. This evidence, combined with the physical localization data for the met locus presented in the accompanying paper, places the CF locus in the middle third of the long arm of chromosome 7, probably between bands q21 and q31.     

Model for antenatal diagnosis of beta-thalassaemia and other monogenic disorders by molecular analysis of linked DNA polymorphisms

Polymorphisms of DNA restriction sites within the human fetal globin genes have been used to identify chromosomes that carry beta-thalassaemia genes in individuals heterozygous for this disease. This has allowed an antenatal diagnosis for beta-thalassaemia to be carried out by observation of the pattern of the inherited polymorphism of a linked DNA sequence not involved in the genetic pathogenesis of the disease. In the populations we have investigated there is no constant pattern of polymorphism that segregates with the beta-thalassaemia gene. The use of linked polymorphisms should, therefore, be applicable to antenatal diagnosis both of beta-thalassaemia and of any other single-gene defect for which there is a DNA probe specific for a sequence linked to the affected locus.     

Clinical and genetic analysis of two Chinese families with benign familial neonatal convulsions

Benignfamilialneonatalconvulsions(BFNC)is arareautosomaldominantinheritedepilepsysyn dromecharacterizedbyunprovokedpartialorgeneral izedseizures.Theseizuresusuallyoccurfromthesec onddayofbirthtothesixthmonthandremitsponta neouslyafterseveralweekstomonths.Mostindivid ualsareseizure freebytheageofsixmonths.The serumchemistryandneuroradiologicalexaminations,interictalelectroencephalogram(EEG),andpsy chomotordevelopmentareusuallynormal.However,10%to15%ofpatientshavetheriskofseizurere currencela…     

Clinical and genetic analysis of two Chinese families with benign familial neonatal convulsions

Benign familial neonatal convulsions (BFNC) is a rare autosomal dominant inherited epilepsy syndrome. Two voltage-gated potassium channel genes, KCNQ2 and KCNQ3, have been identified as the genes responsible for BFNC. Here we report two Chinese families with clinical histories of typical BFNC. Using six microsatellite markers, two located at KCNQ2 locus and four at KCNQ3 locus, linkage analysis was performed in the two families, which excluded the linkage of BFNC to KCNQ3, but could not exclude the linkage to KCNQ2. Direct DNA sequencing of the KCNQ2 gene in the two families was performed, and two formerly unknown polymorphisms were identified, but no KCNQ2 mutation was found in the two families. Our study suggests the genetic heterogeneity in Chinese families with BFNC and proves the existence of a new gene locus for BFNC.     

A frame-shift mutation in the cystic fibrosis gene.

Cystic fibrosis (CF) is a common recessive lethal genetic disorder, affecting 1 in 1,600 Caucasians. The disease causes defective regulation of chloride-ion transport in exocrine cells. Although in all CF families the disease is linked to a locus on chromosome 7q31, there is clinical heterogeneity in the severity of the disease and the age at which it is diagnosed. CF is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. A three-nucleotide deletion (delta F508) causing the loss of a phenylalanine residue in the tenth exon of the CFTR gene has been found on 70% of CF chromosomes. We have now characterized a CF family in which neither parent of the affected individual carries the common mutation, and identified a two-nucleotide insertion in the CF allele of the mother. The mutation introduces a termination codon in exon 13 of the CFTR gene at residue 821, and is predicted to result in the production of a severely truncated nonfunctional protein.     

Assignment of multiple endocrine neoplasia type 2A to chromosome 10 by linkage

Multiple endocrine neoplasis type 2A (MEN2A) is one of several kinds of cancers that appear to be inherited in an autosomally dominant fashion. We have assigned the MEN2A locus to chromosome 10 by linkage with a new DNA marker (D10S5). The linkage led us to investigate other chromosome 10 markers and demonstrate linkage between the disease locus and the interstitial retinol-binding protein (IRBP) gene. The D10S5 locus was sublocalized to 10q21.1 by hybridization in situ and the IRBP gene to p11.2----q11.2 with a secondary site at q24----q25. The linkages were established using 292 members of five families, three different restriction fragment length polymorphisms (RFLPs) at D10S5 and two RFLPs recognized by the IRBP probe. The recombination frequencies from pairwise linkage analysis between the disease and two marker loci D10S5 and IRBP were 0.19 and 0.11, with maximum lod scores of 3.6 and 8.0 respectively. Ordering of the three loci by multipoint analysis placed the IRBP gene approximately midway between the disease and D10S5 loci.     

A linked genetic marker for multiple endocrine neoplasia type 2A on chromosome 10

Multiple endocrine neoplasia type 2A (MEN2A) is an autosomal dominantly inherited cancer syndrome characterized by medullary carcinoma of the thyroid, phaeochromocytoma and hyperparathyroidism. Almost all gene carriers can be detected by screening tests before the age of 40, but the nature and location of the predisposing gene are unknown. Simpson et al. recently reported preliminary evidence for linkage between the DNA probe p9-12A on chromosome 10 and MEN2A. We now report linkage between the MEN2A locus and the interstitial retinol-binding protein gene, which is located on chromosome 10p11.2-q11.2.     

中国榛属植物DNA提取与SSR初步分析

为了探讨适合中国榛属植物基因组DNA的提取方法,分析榛属植物的遗传多样性,本实验以榛属植物的叶片为试材,通过对Doyle和Doyle方法的改良,摸索出适于中国榛属植物基因组DNA提取的方法,并以核酸产量、纯度、片断分布情况等指标来评价,获得高质量基因组DNA;同时应用4对欧榛SSR引物对中国榛属植物进行了跨种转移,并对具有商业潜力的平榛、毛榛和川榛3个种的遗传多样性进行了初步评价,4对引物从3个种的29个样本中扩增出33个等位基因,位点拥有的等位基因数量在6~12个之间,位点平均等位基因数为8.38.上述结果表明SSR是用于榛属种间育种、品种鉴定以及种间遗传图构建的有力工具.     

Expression and characterization of the cystic fibrosis transmembrane conductance regulator.

Cystic fibrosis (CF) is a common lethal genetic disease that manifests itself in airway and other epithelial cells as defective chloride ion absorption and secretion, resulting at least in part from a defect in a cyclic AMP-regulated, outwardly-rectifying Cl- channel in the apical surface. The gene responsible for CF has been identified and predicted to encode a membrane protein termed the CF transmembrane conductance regulator (CFTR). Identification of a cryptic bacterial promoter within the CFTR coding sequence led us to construct a complementary DNA in a low-copy-number plasmid, thereby avoiding the deleterious effects of CFTR expression on Escherischia coli. We have used this cDNA to express CFTR in vitro and in vivo. Here we demonstrate that CFTR is a membrane-associated glycoprotein that can be phosporylated in vitro by cAMP-dependent protein kinase. Polyclonal and monoclonal antibodies directed against distinct domains of the protein immunoprecipitated recombinant CFTR as well as the endogenous CFTR in nonrecombinant T84 cells. Partial proteolysis fingerprinting showed that the recombinant and non-recombinant proteins are indistinguishable. These data, which establish several characteristics of the protein responsible for CF, will now enable CFTR function to be studied and will provide a basis for diagnosis and therapy.     

No evidence for illegitimate young in monogamous and polygynous warblers

In animals with internal fertilization, paternity is uncertain. In birds, the occurrence of copulations outside the pair-bond has been documented in a number of species, but the extent to which these result in illegitimate young is largely unknown, and constitutes a major deficiency in our understanding of avian mating systems. The analysis of tandemly repeated sequences (minisatellites), has enhanced our ability to make individual identifications and paternity determinations. Here we describe the use of a bird minisatellite DNA probe in assigning paternity in natural populations of the monogamous willow warbler Phylloscopus trochilus and of the polygynous wood warbler Phylloscopus sibilatrix. In both species this probe detects a multiple locus pattern and a single locus that exhibits a variable number of tandem repeats. Although we observed intrusions by non-resident males into the territories of paired males and extra-pair copulations, no illegitimate offspring were detected among 176 young from 32 families of both species, implying that extra-pair copulations have little or no genetic impact.     

Abnormal pattern detected in fragile-X patients by pulsed-field gel electrophoresis.

The fragile-X syndrome is the most frequent inherited form of mental retardation, with an incidence of 1 in 1,500 males. It is characterized by the presence of a fragile site at Xq27.3 induced in vitro by folate deprivation or by inhibitors of deoxynucleotide synthesis. Its mode of inheritance is unusual for an X-linked trait, with incomplete penetrance in both males and females. Some phenotypically normal males transmit the mutation to all their daughters who rarely express any symptoms, but penetrance is high in sons and daughters of these carrier women. Genetic and physical mapping of the Xq27-q28 region has confirmed that the disease locus is located at or very near the fragile site. Hypotheses proposed to account for the abnormalities in the inheritance of the disease include sequence rearrangements by meiotic recombination or a mutation that affects reactivation of an inactive X chromosome during differentiation of female germ cells. To detect such rearrangements, or methylation changes that may reflect a locally inactive X chromosome, we used pulsed-field gel analysis of DNA from fragile-X patients with probes close to the fragile-X locus. The probe Do33 (DXS465) detected abnormal patterns in fragile-X patients, but not in normal controls or in non-expressing male transmitters.     

Localization of the malignant hyperthermia susceptibility locus to human chromosome 19q12-13.2

Malignant hyperthermia (MH) is an inherited human skeletal muscle disorder and is one of the main causes of death due to anaesthesia. The reported incidence of MH varies from 1 in 12,000 in children to 1 in 40,000 in adults. MH is triggered in susceptible people by all commonly used inhalational anaesthetics; it is characterized by a profoundly accelerated muscle metabolism, contractures, hyperthermia and tachycardia. Susceptibility to MH (MHS) is predicted by contracture tests on muscle tissue obtained by biopsy. An almost identical disorder known as porcine MH exists in pigs. The genetics of the porcine syndrome have been extensively studied; the locus controlling expression of porcine MH is genetically linked to the glucose phosphate isomerase locus (GPI). In man, GPI has been mapped to the q12-13.2 region of chromosome 19 (refs 10-12). We have now investigated genetic linkage in several extended Irish pedigrees in which MHS is segregating as an autosomal dominant trait. Here we show linkage between MHS and DNA markers from the GPI region of human chromosome 19 with a maximum log likelihood ratio (lod score) of 5.65 at the CYP2A locus. These results indicate that human and porcine MH are most probably due to mutations in homologous genes, and also provide a potentially accurate and noninvasive method of diagnosis for MHS.     

Detection of deletions spanning the Duchenne muscular dystrophy locus using a tightly linked DNA segment

The Duchenne muscular dystrophy (DMD) locus has been localized to the short arm of the human X chromosome (Xp21) by detection of structural abnormalities and by genetic linkage studies. A library highly enriched for human DNA from Xp21 was constructed using DNA isolated from a male patient who had a visible deletion and three X-linked disorders (DMD, retinitis pigmentosa and chronic granulomatous disease). Seven cloned DNA probes from this library and the probe 754 (refs 5, 8) are used in the present study to screen for deletions in the DNA isolated from 57 unrelated males with DMD. Five of these DMD males are shown to exhibit deletions for one of the cloned DNA segments and at least 38 kb of surrounding DNA. In addition, two subclones from the same region detect four restriction fragment length polymorphisms which exhibit no obligate recombination with DMD in 34 meiotic events. These new DNA segments will complement the existing Xp21 probes for use in carrier detection and prenatal diagnosis of DMD. Elucidation of the end points of the five deletions will help delineate the extent of the DMD locus and ultimately lead to an understanding of the specific sequences involved in DMD.     

Regional localization of sex-specific Bkm-related sequences on proximal chromosome 17 of mice

Development and fertility in the mouse are known to be influenced by loci mapped to the T/t complex of chromosome 17. Recent evidence suggests that one or more genes near this region may also be associated with sex determination. Washburn and Eicher recently reported partial to complete sex reversal with the Thp deletion on some genetic backgrounds and suggest that this result may be due to a primary sex-determining locus (Tas) that is closely linked to, or a part of, the T locus. Sex-specific, Bkm (banded Krait minor satellite DNA)-related sequences are known to have autosomal as well as heterogametic sex chromosomal copies, but specific regions of autosomal localization have not been described. We now demonstrate the presence of chromosome Y-related DNA sequences on proximal chromosome 17 in Sex-reversed (Sxr) and normal mice using in situ hybridization of mitotic chromosomes with 3H-labelled pCS316 (ref. 4), a probe that shows major hybridization to the proximal portion of the mouse chromosome Y. These data, and those of Washburn and Eicher, argue for a gene(s) related to sex determination or differentiation within the proximal portion of mouse chromosome 17.     

A highly polymorphic locus very tightly linked to the Huntington's disease gene

The genetic defect in Huntington's disease (HD), an inherited neuropsychiatric disorder of unknown etiology, has not been defined. The discovery of linkage between HD and the DNA marker D4S10(G8) raised the possibility of isolating the disease gene on the basis of its chromosomal location, in addition to providing a limited presymptomatic test for the late onset disorder. But it has been difficult to isolate other DNA markers nearer to the HD gene, and this has hampered attempts to identify the disease locus and limited the applicability and accuracy of predictive testing. Recently, several new DNA markers from the region of the genome near the HD gene have been isolated using a directed cloning strategy. We describe here the characterization of one of these new markers, D4S95, a highly polymorphic locus which displays no recombination with the HD gene in the families tested. The high degree of polymorphism at this locus and its proximity to the HD gene make it extremely useful for predictive testing and as a new starting point for attempts to clone the disease gene.     

The genetic defect in familial Alzheimer's disease is not tightly linked to the amyloid beta-protein gene

Amyloid beta-protein (AP) is a peptide of relative molecular mass (Mr) 42,000 found in the senile plaques, cerebrovascular amyloid deposits, and neurofibrillary tangles of patients with Alzheimer's disease and Down's syndrome (trisomy 21). Recent molecular genetic evidence has indicated that AP is encoded as part of a larger protein by a gene on chromosome 21 (refs 5-7). The defect in the inherited autosomal dominant form of Alzheimer's disease, familial Alzheimer's disease (FAD), has been mapped to the same approximate region of chromosome 21 by genetic linkage to anonymous DNA markers, raising the possibility that this gene product, which could be important in the pathogenesis of Alzheimer's disease, is also the site of the inherited defect in FAD (ref. 5). We have determined the pattern of segregation of the AP gene in FAD pedigrees using restriction fragment length polymorphisms. The detection of several recombination events with FAD suggests that the AP gene is not the site of the inherited defect underlying this disorder.     

Plant genetics: RNA cache or genome trash?

According to classical mendelian genetics, individuals homozygous for an allele always breed true. Lolle et al. report a pattern of non-mendelian inheritance in the hothead (hth) mutant of Arabidopsis thaliana, in which a plant homozygous at a particular locus upon self-crossing produces progeny that are 10% heterozygous; they claim that this is the result of the emerging allele having been reintroduced into the chromosome from a cache of RNA inherited from a previous generation. Here I suggest that these results are equally compatible with a gene conversion that occurred through the use as a template of DNA fragments that were inherited from a previous generation and propagated in archival form in the meristem cells that generate the plant germ lines. This alternative model is compatible with several important observations by Lolle et al..