Primer-engineered multiplex PCR–RFLP for detection of MTHFR C677T, prothrombin G20210A and factor V Leiden mutations

Köksal, Vedat; Barış, İbrahim; Etlik, Özdal

doi:10.1016/j.yexmp.2006.12.006

Cited by 22 publications

(18 citation statements)

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“…It has been shown that homozygosity for the C to T substitution at nucleo tide 677 of the MTHFR gene is associated with a 30-50 % reduction of this enzyme activity and is the most common inherited cause of moderate hyperhomocysteinemia [1,2]. In the last publica tions there are evidences that the C677T MTHFR gene variation is a risk factor for ischemic stroke [2], infertility [1], primary closed angle glaucoma (PCAG) [3], presence of anti hepatitis B virus (HBV) antibodies [4], cytomegalovirus infection (CMV) [4], Huntington disease [5], cancer [6], coronary artery disease (CAD) [7], neural tube defects (spina bifida) and venous thrombosis [6,8,9], but the association of the C677T MTHFR gene variation and the risk of primary open angle glau coma (POAG) is still conflicting [3].…”

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confidence: 99%

“…APC is a serine protease with potent anti coagulant properties, which is formed in blood on the endothelium from an inactive precursor [10]. The point mutation of factor V (guanine is replaced by adenine in position 1691) led to the structural change in factor V molecule (F5 Q506, or FV Leiden) that is not properly inactivated by APC and shifts the balance toward thrombosis in the clotting cas cade [2,[9][10][11][12][13][14][15] and may increase the risk of sys temic lupus erythematosus (SLE) [12], type 2 dia betes [16], recurrent pregnancy loss (RPL) [17][18][19][20][21], stroke [2,22], venous thrombosis and myocar dial infraction [2,22].…”

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confidence: 99%

“…It has been shown that F2 gene G20210A SNP is associated with higher plasma prothrombin con centrations and augmented thrombin generation [11,17,20,23,24]. Patients carrying the 20210A allele have been demonstrated to be at risk for venous thrombosis [9,11,15,17,20], RPL and ischemic stroke [18][19][20].…”

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Allelic polymorphism of F2, F5 and MTHFR genes in population of Ukraine

2010

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Allelic polymorphism of F2, F5 and MTHFR genes in population of Ukraine

2010

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“…On the other hand, 677CT genotype has MTHFR enzyme activity of about 65% when compared to wild type (Curtin et.al, 2004). The substitution of "T" for "C" at 677 position of MTHFR gene creates a restriction site for the enzyme Hind111 (Koksal et al, 2007). During electrophoresis of the PCR products 25bp fragment migrates out of the gel due to the small fragment size.…”

Section: Discussionmentioning

confidence: 99%

Investigation of Methyl Tetrahydro-Folate Reductase C677t Polymorphism in a Sample of Patients in a Hospital Based Diagnostic Setting in Sri Lanka

Varnakulasingham¹,

Manamperi²

2016

International Conference on Bioscience and Biotechnology

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The present study was conducted to identify the prevalence of MTHFR gene mutation in a group of Sri Lankan patients seeking routine diagnostic testing for the C677T mutation through PCR followed by RFLP diagnostic methods. Human MTHFR gene codes for the production of MTHFR enzyme in humans which plays major role in homocysteine and folate metabolism in humans. The mutation in MTHFR gene at nucleotide position 677 has two possibilities: C (cytosine, which occurs in the wild type gene) or T (thymine, which occurs in the mutated gene).In C677T variant, modification from C to T gives rise to the replacement of alanine by valine residue in protein at 222 aa position. Eight serum samples were collected from the MTHFR mutation suspected patients. Genomic DNA extraction was done and extracted DNA was subjected to PCR amplification. PCR products were subjected to RFLP assay with overnight digestion. The substitution of "C" to "T" at 677 position of MTHFR gene creates a restriction site for the enzyme Hind111. RFLP products were subjected to gel electrophoresis and further analysis. The results obtained shows that, of the 8 samples tested, 2 had 677CT genotype, 1 had 677TT genotype whereas the remaining 5 had 677CC wild type. Homozygous 677TT genotype leads to lower MTHFR activity and hyperhomocysteinaemia, which can cause defects in DNA repair. However, as this is a small sample of the population, it is recommended to carry out studies with a large sample volume to ascertain the prevalence of this mutation in the community.

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“…Angelini, et al, identificaron estos tres factores de riesgo genético empleando PCR múltiple específica de alelo y variando el método de obtención de ADN en busca de una técnica rápida, fácil y económica (70). En el 2007, Koksal y su grupo presentaron una modificación en los cebadores empleados para la amplificación simultánea del FV de Leiden, metilen-tetrahidrofolato reductasa C677T y protrombina G20210A (PCR múltiple) con el fin de crear un sitio de restricción para la enzima de digestión (Mnl I) y lograr un método más rápido, fácil y económico que se pudiera implementar en la determinación rutinaria de laboratorio (71).…”

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Genotipos frecuentemente asociados a trombofilias

2013

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Contribución de los autores:Solangy Usme: revisión y recopilación de la información. Helena Hernández-Cuervo: revisión de la información, escritura y edición del documento. Juan José Yunis: dirección del grupo de investigación y revisión del escrito. La enfermedad tromboembólica venosa es una entidad patológica importante debido a la morbilidad que causa, por sus complicaciones y por su alta incidencia en el mundo, la cual puede variar desde 1:100 en adultos mayores hasta 1:100.000 en niños. Existen múltiples factores de riesgo tanto genéticos como ambientales asociados a la enfermedad; los más ampliamente estudiados por su incidencia en la población mundial son el factor V de Leiden, la mutación G20210A en el factor II (protrombina) y las mutaciones en la metilen-tetrahidrofolato reductasa C677T y A1298C, que hasta hace poco se consideraban factores de riesgo. En la presente revisión se presenta de forma concisa qué es y cómo se produce un trombo a partir de la enfermedad tromboembólica, cuáles son las principales entidades nosológicas que involucra la enfermedad y los genotipos más frecuentemente asociados a la misma. Se enfatiza en las alteraciones genéticas epidemiológicamente más importantes y se muestran brevemente los estudios realizados en Colombia.Palabras clave: trombofilia, protrombina, trombosis, trombosis de la vena, embolia pulmonar. doi: http://dx.doi.org/10.7705/biomedica.v34i1.839 Frequently associated genotypes to thrombophiliaVenous thromboembolism is an important pathological entity that causes high morbidity due either to the disease or its complications. The incidence in the world ranges between 1:100 in adults and 1:100,000 in children. Risk factors for the disease include genetic as well as environmental factors. Among them, factor V Leiden (G1691A), prothrombin (G20210A) and MTHFR C677T and A1298C (which until recently were considered risk factors), have been widely studied given their impact in the world. This review presents in a clear and concise way what a thrombous is and how it is formed; how a clot is able to produce thromboembolic disease; what are the main nosological entities involved, and their main genetic causes. The most epidemiologically important genetic alterations and studies conducted in Colombia will be emphasized. Las alteraciones de los procesos de coagulación pueden conllevar a cualquiera de dos estados: 1) hipercoagulabilidad o trombofilia (predisposición excesiva a la formación de trombos), o 2) hipocoagulabilidad (que aumenta el riesgo de hemorragia).Cualquier alteración o lesión en la pared de los vasos sanguíneos genera la activación de los procesos de coagulación en los que intervienen vías y factores con el fin de mantener la hemostasis y evitar la hemorragia; este proceso ocurre en tres pasos básicos que son la vasoconstricción local, la agregación plaquetaria y la formación del coágulo. La visión actual de la coagulación

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Primer-engineered multiplex PCR–RFLP for detection of MTHFR C677T, prothrombin G20210A and factor V Leiden mutations

Cited by 22 publications

References 14 publications

Allelic polymorphism of F2, F5 and MTHFR genes in population of Ukraine

Allelic polymorphism of F2, F5 and MTHFR genes in population of Ukraine

Investigation of Methyl Tetrahydro-Folate Reductase C677t Polymorphism in a Sample of Patients in a Hospital Based Diagnostic Setting in Sri Lanka

Genotipos frecuentemente asociados a trombofilias

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