Derivation of clones from the choroideremia locus by preparative field inversion gel electrophoresis

By making use of preparative field inversion gel electrophoresis, we have constructed a lambda ZAP library that is highly enriched for sequences from the choroideremia locus. In vivo excision of pBluescript SK(-) constructs from lambda ZAP obviates the subcloning of DNA inserts and allows for rapid...

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Published inNucleic acids research Vol. 18; no. 4; pp. 725 - 731
Main Authors VAN DE POL, T. J. R, CREMERS, F. P. M, BROHET, R. M, WIERINGA, B, ROPERS, H.-H
Format Journal Article
LanguageEnglish
Published Oxford Oxford University Press 25.02.1990
Subjects
Animals
Bacteriophage lambda - genetics
Biological and medical sciences
choroideremia
Choroideremia - genetics
Chromosome Mapping
Cloning, Molecular
Cricetinae
Cricetulus
Deoxyribonuclease EcoRI
DNA - genetics
DNA - isolation & purification
Electrophoresis, Agar Gel - methods
Escherichia coli - genetics
Fundamental and applied biological sciences. Psychology
Genes. Genome
Humans
Hybrid Cells - cytology
linkage analysis
Male
Molecular and cellular biology
Molecular genetics
Polymorphism, Restriction Fragment Length
Reference Values
Restriction Mapping
X Chromosome
Genetic mapping
Human
Pulsed field electrophoresis
Restriction fragment length polymorphism
Gene
Choroideremia
Chromosome DNA
Gene library
Molecular cloning
Chorioretinopathy
Genetic disease
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Summary:By making use of preparative field inversion gel electrophoresis, we have constructed a lambda ZAP library that is highly enriched for sequences from the choroideremia locus. In vivo excision of pBluescript SK(-) constructs from lambda ZAP obviates the subcloning of DNA inserts and allows for rapid processing of several hundred recombinants. From a 625 kb Sfil fragment we isolated 7 clones that were physically mapped using microdeletions associated with the disease. One of these clones is located within, or just telomeric to, the choroideremia gene and detects two restriction fragment length polymorphisms (RFLPs). Another clone detects a RFLP which maps centromeric to the disease locus. Together these probes should improve the reliability of linkage analysis in choroideremia families and should pave the way for the isolation of the choroideremia gene.
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ISSN:0305-1048
1362-4962
DOI:10.1093/nar/18.4.725