Isolation of probes detecting restriction fragment length polymorphisms from X chromosome-specific libraries: potential use for diagnosis of Duchenne muscular dystrophy

Hofker, Marten H.; Wapenaar, Martin C.; Goor, Nicole; Bakker, Egbert; Ommen, G.J.B. van; Pearson, P. L.

doi:10.1007/bf00273073

Cited by 191 publications

(33 citation statements)

References 27 publications

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“…DNA was isolated from approximately 2 x 10 s trypsinized cells by the isopropanol precipitation method described elsewhere (Hofker et al 1985} and dissolved in 10 mM Tris, 1 mM EDTA (pH 8) to a final concentration of 300 }zg/ml. The DNA was then restricted with EcoRI or, in the case of the L1 repeat analysis, with HindIII, and the fragments were separated according to size by electrophoresis through a 0.8% agarose gel.…”

Section: Southern Analysismentioning

confidence: 99%

Investigation of chromosome-mediated gene transfer using the HPRT region of the human X chromosome as a model.

Pritchard¹,

Goodfellow²

1987

Genes Dev.

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A panel of over 50 hybrid cells containing varying portions of the long arm of the human X chromosome have been obtained by chromosome-mediated gene transfer (CMGT) of human chromosomes to mouse cells deficient in HPRT. This panel is used to investigate the size and integrity of transfected human chromosome fragments and also to examine the effect of including a selectable DNA plasmid in the transfection mix. Chromosomal rearrangements are found to be generated in the chromosome transfer process, and the human X centromeric re#on is detected in the transfected cells at an unusually high frequency. Extensive lengths of X chromosome DNA are transferred intact, suggesting potential uses of CMGT in cloning large genes and loci for which only the chromosomal map position is known.[Key Words: Chromosome transfer; human gene mapping; X chromosome; centromere selection] Received November 6, 1986; revised version received and accepted January 5, 1987.Chromosome-mediated gene transfer (CMGT) can be used to construct hybrid cells that retain fragments of the selected donor chromosome (Mcbride and Ozer 1973}. Chromosome breakage is generally assumed to be random and the chromosome fragment size can vary from 50 kb up to a whole chromosome (Wullems et al. 1976;Olsen et al. 1981). The intermediate size of the transgenome produced by CMGT provides a method for linking and ordering genes over longer distances than the high-resolution mapping techniques of DNA-mediated gene transfer or cosmid cloning and at a more sensitive level than can be achieved by somatic cell hybrids or in situ hybridization. In addition, if fragmentation is random, the frequency with which loci are separated by CMGT could be used as a measure of genetic distance (for review, see Mcbride and Peterson 1980). Despite these advantages CMGT has been restricted in its applications. One major drawback has been the lack of suitable selective methods for isolating the cells containing the required chromosomal fragment. Until recently, only those regions of the genome encoding endogenous selectable loci were amenable to analysis, the best examples being the HPRT locus on the long arm of the human X chromosome (Miller and Ruddle 1978; and the TK locus on the long arm of human chromosome 17 (Klobutcher and Ruddle 1979).In previous experiments, new selection methods were used to extend the number of regions of the genome that could be subjected to analysis by CMGT (Pritchard and Goodfellow 1986). By cotransferring the selectable DNA plasmid, pSV2neo, with human chromosomes to mouse HPRT-cells, it was shown that the frequency of combined uptake of the HPRT chromosomal locus with pSV2neo was close to the frequency of uptake of HPRT alone. This observation demonstrated the feasibility of including an unlinked selectable marker in the transfection mix, followed by preselection for incorporation of that marker as an enrichment step in CMGT experiments. Furthermore, the fact that the neo gene and the HPRT chromosomal locus remained associated after a second chromosome tran...

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Section: Southern Analysismentioning

confidence: 99%

Investigation of chromosome-mediated gene transfer using the HPRT region of the human X chromosome as a model.

Pritchard¹,

Goodfellow²

1987

Genes Dev.

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“…(20). Genomic DNA samples from an individual with normal hemoglobin (HbA) alleles was prepared as described (21). Genomic DNA samples containing hemoglobin sickle (HbS) and hemoglobin C (HbC) alleles were provided by D. Higgs (Oxford) and H. Kazazian (Baltimore).…”

mentioning

confidence: 99%

Attachment of a 40-base-pair G + C-rich sequence (GC-clamp) to genomic DNA fragments by the polymerase chain reaction results in improved detection of single-base changes.

Sheffield

Cox

Lerman

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

1,098

487

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Denaturing gradient gel electrophoresis (DGGE) can be used to distinguih two DNA molecules that differ by as little as a single-base substitution. This method detects =50% of all possible single-base changes in DNA fragments ranging from 50 to -1000 base pairs. To increase the number of single-base changes that can be distingished by DGGE, we used the polymerase chain reaction to attach a 40-base-pair G+C-rich sequence, designated a GC-clamp, to one end of amplified DNA fragments that encompass regions of the mouse and human 13-globin genes. We show that this GC-clamp allows the detection of mutations, includig the hemoglobin sickle (HbS) and hemoglobin C (HbC) mutations within the human P-globin gene, that were previously indiishable by DGGE. In addition to providing an easy way to attach a GC-clamp to genomic DNA fragments, the polymerase chain reaction technique greatly increases the sensitivity of DGGE. With this approach, DNA fragments derived from <5 ng of human genomic DNA can be detected by ethidium bromide staining of the gel, obviating the need for radioactive probes. These improvements extend the applicability of DGGE for the detection of polymorphisms and mutations in genomic and cloned DNA.Genetic linkage analysis has been severely limited in humans due to a paucity of informative polymorphic loci. The recognition that DNA sequence polymorphism can fill this void has revolutionized the field of human genetics (1, 2). Consequently, approaches that allow the detection of singlebase differences in specific regions of genomic DNA have been extremely powerful for both linkage analysis and direct detection of mutations associated with human disease (3-9). The vast majority of base changes have been identified by the restriction fragment length polymorphism approach, which measures DNA sequence alterations due to a loss or gain of a restriction enzyme cleavage site or to variation in length caused by deletion or insertion (1). However, many singlebase changes do not alter a restriction enzyme cleavage site and, therefore, cannot be detected by the restriction fragment length polymorphism method. One alternative method that makes it possible to detect a larger fraction of all possible base changes in a DNA fragment is denaturing gradient gel electrophoresis (DGGE;.DGGE is a gel system that separates DNA fragments according to their melting properties (10-12). When a DNA fragment is electrophoresed through a linearly increasing gradient of denaturants, the fragment remains double stranded until it reaches the concentration of denaturants equivalent to a melting temperature (tm) that causes the lower-temperature melting domains of the fragment to melt. At this point, the branching of the molecule caused by partial melting sharply decreases the mobility of the fragment in the gel. The lower-temperature melting domains of DNA fragments differing by as little as a single-base substitution will melt at slightly different denaturant concentrations because ofdifferences in stacking interactions between adja...

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“…No new alleles were found. Among the observed alleles, those which have previously been reported to be less common [Davies et al, 1983: Murray et al, 1982Aldridge et al, 1984;Hofker et al, 1985] were also rela tively rare in both populations studied here. The allele frequencies are shown in table 11.…”

Section: Resultsmentioning

confidence: 92%

“…The frequencies of the allelic restriction fragments associated with L1.28, RC8, pD2, and 754 were first described for resi dent Dutch and British populations and North American whites, respectively [Davies et al, 1983;Murray et al, 1982;Aldridge et al, 1984;Hofker et al, 1985] (table III). Furthermore, numerous re ports on family studies in different popu lations have documented that these poly morphisms are widely distributed at least among various European populations [e.g., see Wieacker et al, 1983 a, b].…”

Section: Resultsmentioning

confidence: 99%

Four DNA Polymorphisms on the Short Arm of the X Chromosome: Allele Frequencies in a German and in a Turkish Population

1987

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Four restriction fragment length polymorphisms, revealed by cloned arbitrary X chromosome segments (L1.28, RC8, pD2, 754) were studied in samples (50 individuals each) of a German and a Turkish population. All previously reported alleles of these polymorphisms were found in both populations, except the infrequent RC8 allele B3 (3.0 kb fragment), which was absent in both groups. The observed minor alleles were found to be rarer in the German series than in the Turkish group, but there was no conclusive evidence of essentially different allele frequencies in either population. However, the frequencies of the RC8 allele B2 (5.3 kb fragment) were differing at the 5% significance level. The allele frequencies of the four polymorphisms are presented and compared with those reported from other European regions.

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Isolation of probes detecting restriction fragment length polymorphisms from X chromosome-specific libraries: potential use for diagnosis of Duchenne muscular dystrophy

Cited by 191 publications

References 27 publications

Investigation of chromosome-mediated gene transfer using the HPRT region of the human X chromosome as a model.

Investigation of chromosome-mediated gene transfer using the HPRT region of the human X chromosome as a model.

Attachment of a 40-base-pair G + C-rich sequence (GC-clamp) to genomic DNA fragments by the polymerase chain reaction results in improved detection of single-base changes.

Four DNA Polymorphisms on the Short Arm of the X Chromosome: Allele Frequencies in a German and in a Turkish Population

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