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An alternative strategy for identification of type IIA VWD mutations
Author(s) -
Winter P. C.,
Mayne E. E.
Publication year - 1993
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.1993.tb03110.x
Subject(s) - pseudogene , genetics , point mutation , gene , dna , restriction site , microbiology and biotechnology , biology , mutation , plasmid , nucleotide , dna sequencing , restriction enzyme , genome
Summary. A 306 base pair segment of the von Willebrands factor (VWF) gene between nucleotides 4627 and 4932 was amplified by PCR using DNA from a type IIA VWD patient. The amplified DNA was cloned in the plasmid pCRII. Clones of the VWF gene and pseudogene were distinguished by their KpnI restriction patterns. A series of six VWF gene clones was sequenced. A single C to T point mutation at nucleotide 4789 resulting in the substitution of arginine 834 by tryptophan was identified in four clones. This mutation, which destroys a BstEII restriction site, was also detected in amplified DNA of affected relatives of the patient. The method described avoids difficulties associated with strategies employed previously for identification of VWD mutations.