Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This study compared a rapid method to detect the nucleotide mutation 1691 G-->A, responsible for the factor V Arg506-->Gln substitution, with a previously established denaturing gradient gel electrophoresis (DGGE) technique in 136 patients with unexplained thrombosis. The new method comprises amplification of the factor V gene exon 10 with a modified oligonucleotide, permitting the introduction of a cleavage site for the restriction endonuclease HindIII in the fragments bearing the mutation. This simple, rapid, inexpensive and nonisotopic method gave the same results as the DGGE method in all subjects tested.
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PMID:A rapid screening method for the factor V Arg506-->Gln mutation. 765 39

Resistance to the anticoagulant effects of activated protein C (APC) is now considered the most prevalent cause of inherited thrombophilia. The great majority of patients with activated protein C resistance (APCR) have a missense mutation in the factor V molecule (factor V Leiden, FVR506Q) resulting in defective inactivation of factor Va due to a loss of an APC cleavage site. The diagnosis of APCR has been based upon the inability of APC to prolong the activated partial thromboplastin (aPTT) clotting time in subjects with APCR. However, this assay has a number of deficiencies which limit its general use. We have evaluated a newly described one-stage tissue factor dependent factor V coagulation assay for APCR in 117 patients and controls and compared the results of this assay in a blinded manner to a polymerase chain reaction (PCR) based assay for the molecular defect of factor V Leiden. 43% (50/117) of the patients studied were receiving coumadin or heparin, or had a lupus anticoagulant. The tissue factor dependent factor V assay had 100% specificity and sensitivity for factor V Leiden and successfully predicted a homozygous state in the three documented homozygotes. The PCR-based assay for factor V Leiden resulted in a single false positive assay due to a silent A to C transition at nucleotide 1692 resulting in the loss of the Mnl restriction endonuclease cleavage site. The single-stage tissue factor dependent factor V assay is a highly sensitive and generally applicable assay for APCR.
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PMID:Evaluation of a tissue factor dependent factor V assay to detect factor V Leiden: demonstration of high sensitivity and specificity for a generally applicable assay for activated protein C resistance. 894

The factor V Leiden mutation, a G-->A transition at position 1691 in exon 10 of the gene that codes for factor V, produces an Arg506Gln substitution and is the most common genetic risk factor for venous thrombosis. We have developed a rapid, sensitive, and specific method to detect the factor V Leiden mutation in genomic DNA from whole blood by PCR amplification and microparticle enzyme immunoassay detection using the Abbott LCx instrument. We compared this automated method with the standard procedure using restriction endonuclease digestion of PCR products followed by gel electrophoresis in blinded experiments. In 130 patients (from Veterans Affairs medical centers) with deep venous thromboses, including 24 heterozygotes with the factor V Leiden mutation, there was complete agreement between the two methods. The assay was also able to distinguish heterozygotes from homozygotes. This method, which carries a low potential for cross-contamination of samples, should be a useful routine test for the factor V Leiden mutation in clinical laboratories with sufficient demand for molecular diagnostic assays using the LCx instrument.
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PMID:Automated detection of the factor V Leiden mutation using the LCx microparticle enzyme immunoassay. 989 36

Several clinical and environmental conditions are causally related to sudden death from acute pulmonary thromboembolism (APT). Morbid obesity, despite its frequency and association with adverse health effects, is usually considered at most only an additive risk factor for APT. We reviewed protocols and histories from 7227 consecutive autopsies performed between 1985 and 1996 at the Mayo Clinic, including all deaths from APT where no clinical or environmental risk factor could be identified in the study. Body mass indices (BMI) were calculated and compared with those of age- and sex-matched controls who had died suddenly and naturally without evidence of APT. Resistance to activated protein C is the most common molecular clotting defect predisposing to APT, and it is caused by a point mutation in the factor V gene (R506Q). Genomic DNA was extracted from archival tissues of all cases and controls, and the R506Q status was determined by polymerase chain reaction amplification, restriction endonuclease digestion, and direct sequencing. APT was found as the immediate cause of death in 433 patients, with 36 (8%) having no previously established risk factors. Twenty-four of these persons (67%) were morbidly obese (BMI >30 kg/m2). compared with only five controls (14%, P<0.0001). Four patients in both groups, each with a BMI <30 kg/m2. had at least one allele positive for R506Q. Morbid obesity is an independent risk factor in cases of sudden death from APT after the exclusion of previously established clinical, environmental, and molecular risk factors.
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PMID:Death from pulmonary thromboembolism in severe obesity: lack of association with established genetic and clinical risk factors. 1039 88

The G1691A (Leiden) mutation of the factor V gene is the most prevalent identified cause of venous thrombosis. Therefore, we developed a new genetic test using the TaqMan system. With this assay which combines PCR amplification and detection reaction in one closed tube, a cohort of 234 patients with a history of thrombosis was screened. In parallel, amplification products of the same patients were screened with a previously described test using endonuclease digestion of PCR products followed by gel electrophoresis. Identical results were obtained by both methods. Among cases, 122 (52%) individuals were homozygous normal, 99 (42%) were heterozygous affected and 13 (5.5%) showed homozygous pattern for the Factor V Leiden mutation. Thus, it could be demonstrated that the new TaqMan assay is a robust, rapid and automated method for high throughput application which avoids time consuming and difficult post-PCR steps.
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PMID:Allelic discrimination of factor V Leiden using a 5' nuclease assay. 1054 16

Genetic abnormalities in hemostatic proteins associated with hypercoagulability are an important hereditary risk factor for venous thrombosis. Several genetic mutations that cause hereditary disorders predisposing to thrombosis have been described, point mutation in the coagulation factor V gene (FV:R506Q), called factor V Leiden, being the most common of them. A new inexpensive and simple polymerase chain reaction-single-strand polymorphism (PCR-SSCP) based method for detection of this genetic abnormality is reported. The study population consisted of 150 subjects whose factor V genotype was previously determined by PCR-RFLP method using the Mnl I restriction endonuclease. A 223-bp fragment containing the G1692-A (Arg 506-Gln) polymorphic site in exon 10 of the factor V gene was amplified, denatured, and run overnight on the commercially available GMA gels for SSCP. PCR-SSCP analysis showed reproducible and uniform band patterns for FV mutant and wild type alleles. Furthermore, PCR-SSCP results were consistent with those obtained with PCR-RFLP analysis (100%). The described PCR-SSCP procedure is reliable, time-saving, and cost-effective. The method may be considered as a potentially powerful new tool in the routine detection of factor V Leiden.
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PMID:Detection of factor V Leiden by PCR-SSCP using GMA precast Elchrom scientific gels. 1450 11