IDENTIFYING THE C677T POLYMORPHISM OF MTHFR GENE BY PCR-RFLP TECHNIQUE

2011 ◽  
pp. 28-35
Author(s):  
Keyword(s):  
Internal Control ◽  
Restriction Site ◽  
Mthfr Gene ◽  
C677t Polymorphism ◽  
Mthfr Genotype ◽  
Restriction Digest ◽  
Genotype Frequencies ◽  
Pcr Rflp ◽  
Sequencing Technique ◽  
Volunteer Group

Background: The C677T polymorphism of MTHFR gene is a risk factor of many diseases. This study is aimed at: (1) Improving a PCR-RFLP process with the own designed primers to identify the C677T polymorphism of MTHFR gene. (2) Evaluating the prevalence of the C677T polymorphism of MTHFR gene in volunteer group. Materials and method: DNA samples was extracted from peripheral blood of 60 volunteers. Designing primers by using FastPCR software, then improving PCR technique. Standardizing the optimal conditions of restriction digest by HinfI. Confirming the results of polymorphism by DNA sequencing technique. Results: We designed successfully primers to amplify fragment of MTHFR gene including C677T polymorphism and an obligatory restriction site of HinfI (as internal control). 0.5 µl of HinfI enzyme (10 U/µl) is enough for restriction digest. The MTHFR genotype frequencies were: 71.67 % (677CC); 25% (677CT); and 3.33 % (677TT). Conclusion: We standardized successfully PCR-RFLP technique to identifying C677T polymorphism of MTHFR gene. Keywords: C677T polymorphism, MTHFR gene, PCR-RFLP

STUDY ON IDENTIFYING THE C677T POLYMORPHISM OF MTHFR GENE BY PCR-RFLP TECHNIQUE IN PATIENTS WITH ABNORMAL SEMEN ANALYSIS

2012 ◽  
pp. 66-73
Author(s):  
Thi Minh Thi Ha ◽  
Thi Nguyet Minh Nguyen
Keyword(s):  
Internal Control ◽  
Restriction Site ◽  
Semen Analysis ◽  
Mthfr Gene ◽  
C677t Polymorphism ◽  
Mthfr Genotype ◽  
Genotype Frequencies ◽  
Pcr Rflp ◽  
Sequencing Technique ◽  
Volunteer Group

Background: The C677T polymorphism of MTHFR gene is a risk factor of many diseases, including male infertility. This study is aimed at: 1. Improving a PCR-RFLP process with the own designed primers to identify the C677T polymorphism of MTHFR gene. 2. Evaluating the prevalence of the C677T polymorphism of MTHFR gene in the volunteer group and the male with abnormal semen analysis. Materials and Method: DNA samples was extracted from peripheral blood of 60 volunteers and 30 patients with abnormal semen analysis. Designing primers by using FastPCR software, then improving PCR-RFLP technique. Confirming the results of polymorphism by DNA sequencing technique. Evaluating the prevalence of the C677T polymorphism of MTHFR gene. Results: We designed successfully primers to amplify fragment of MTHFR gene including C677T polymorphism and an obligatory restriction site of HinfI (as internal control). The MTHFR genotype frequencies in the volunteer group were: 71.67% (677CC); 25% (677CT); and 3.33% (677TT). Genotype frequencies in patients with abnormal semen analysis were 66.67% ; 30% and 3.33% ; in group of azoospermia were 53.34% ; 40% and 6.66% ; in group of oligospermia were 80%; 20% and 0%. Conclusion: We standardized successfully PCR-RFLP technique to identifying C677T polymorphism of MTHFR gene.We evaluated the prevalence of the C677T polymorphism of MTHFR gene in the volunteer group and the male with abnormal semen analysis.

scholarly journals MTHFR Gene C677T Polymorphism in Autism Spectrum Disorders

2014 ◽  
Vol 2014 ◽  
pp. 1-5 ◽  
Author(s):  
Elif Funda Sener ◽  
Didem Behice Oztop ◽  
Yusuf Ozkul
Keyword(s):  
Autism Spectrum Disorders ◽  
Neurodevelopmental Disorder ◽  
Autism Spectrum ◽  
Mthfr Gene ◽  
C677t Polymorphism ◽  
Autistic Children ◽  
Dsm V ◽  
Pcr Rflp ◽  
Spectrum Disorders ◽  
Genetic And Environmental Factors

Aim. Autism is a subgroup of autism spectrum disorders, classified as a heterogeneous neurodevelopmental disorder and symptoms occur in the first three years of life. The etiology of autism is largely unknown, but it has been accepted that genetic and environmental factors may both be responsible for the disease. Recent studies have revealed that the genes involved in the folate/homocysteine pathway may be risk factors for autistic children. In particular, C677T polymorphism in the MTHFR gene as a possible risk factor for autism is still controversial. We aimed to investigate the possible effect of C677T polymorphism in a Turkish cohort. Methods. Autism patients were diagnosed by child psychiatrists according to DSM-IV and DSM-V criteria. A total of 98 children diagnosed as autistic and 70 age and sex-matched children who are nonautistic were tested for C677T polymorphism. This polymorphism was studied by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods. Results. MTHFR 677T-allele frequency was found to be higher in autistic children compared with nonautistic children (29% versus 24%), but it was not found statistically significant. Conclusions. We conclude that other MTHFR polymorphisms such as A1298C or other folate/homocysteine pathway genes may be studied to show their possible role in autism.

Five genetic polymorphisms of cytochrome P450 enzymes in the Czech non-Roma and Czech Roma population samples

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Lucie Dlouhá ◽  
Věra Adámková ◽  
Lenka Šedová ◽  
Věra Olišarová ◽  
Jaroslav A. Hubáček ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Genetic Polymorphisms ◽  
Metabolic Pathways ◽  
Cytochromes P450 ◽  
Taqman Assay ◽  
Cytochrome P450 Enzymes ◽  
Roma Population ◽  
Czech Population ◽  
Genotype Frequencies ◽  
Pcr Rflp

AbstractObjectivesCytochromes P450 play a role in human drugs metabolic pathways and their genes are among the most variable in humans. The aim of this study was to analyze genotype frequencies of five common polymorphisms of cytochromes P450 in Roma/Gypsy and Czech (non-Roma) population samples with Czech origin.MethodsRoma/Gypsy (n=302) and Czech subjects (n=298) were genotyped for CYP1A2 (rs762551), CYP2A6 (rs4105144), CYP2B6 (rs3745274) and CYP2D6 (rs3892097; rs1065852) polymorphisms using PCR-RFLP or Taqman assay.ResultsWe found significant allelic/genotype differences between ethnics in three genes. For rs3745274 polymorphism, there was increased frequency of T allele carriers in Roma in comparison with Czech population (53.1 vs. 43.7%; p=0.02). For rs4105144 (CYP2A6) there was higher frequency of T allele carriers in Roma in comparison with Czech population (68.7 vs. 49.8%; p<0.0001). For rs3892097 (CYP2D6) there was more carriers of the A allele between Roma in comparison with Czech population (39.2 vs. 38.2%; p=0.048). Genotype/allelic frequencies of CYP2D6 (rs1065852) and CYP1A2 (rs762551) variants did not significantly differ between the ethnics.ConclusionsThere were significant differences in allelic/genotype frequencies of some, but not all cytochromes P450 polymorphisms between the Czech Roma/Gypsies and Czech non-Roma subjects.

scholarly journals Association of Genetic Polymorphisms and Serum Levels of IL-6 and IL-8 with the Prognosis in Children with Neuroblastoma

Cancers ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 529
Author(s):  
Silvia Selene Moreno-Guerrero ◽  
Arturo Ramírez-Pacheco ◽  
Luz María Rocha-Ramírez ◽  
Gabriela Hernández-Pliego ◽  
Pilar Eguía-Aguilar ◽  
...  
Keyword(s):  
Overall Survival ◽  
Genetic Polymorphisms ◽  
Poor Prognosis ◽  
Serum Levels ◽  
Control Group ◽  
Mexican Children ◽  
Genotype Frequencies ◽  
Pcr Rflp ◽  
The Impact ◽  
Circulating Levels

There is evidence that high circulating levels of IL-6 and IL-8 are markers of a poor prognosis in various types of cancer, including NB. The participation of these cytokines in the tumor microenvironment has been described to promote progression and metastasis. Our objective was to evaluate the prognostic role of genetic polymorphisms and serum levels of IL-6 and IL-8 in a cohort of Mexican pediatric patients with NB. The detection of the SNPs rs1800795 IL-6 and rs4073 and rs2227306 IL-8 was carried out by PCR-RFLP and the levels of cytokines were determined by the ELISA method. We found elevated circulating levels of IL-8 and IL-6 in NB patients compared to the control group. The genotype frequencies of the rs1800795 IL-6 and rs4073 IL-8 variants were different between the patients with NB and the control group. Likewise, the survival analysis showed that the GG genotypes of rs1800795 IL-6 (p = 0.014) and AA genotypes of rs4073 IL-8 (p = 0.002), as well as high levels of IL-6 (p = 0.009) and IL-8 (p = 0.046), were associated with lower overall survival. We confirmed the impact on an adverse prognosis in a multivariate model. This study suggests that the SNPs rs1800795 IL-6 and rs4073 IL-8 and their serum levels could be promising biomarkers of a poor prognosis, associated with overall survival, metastasis, and a high risk in Mexican children with NB.

scholarly journals Evaluation of Frequency of PGC1-α and CKMM Genes Polymorphisms Among Iranian Elite Hockey Athletes

2021 ◽  
Vol In Press (In Press) ◽  
Author(s):  
Maryam Valipour ◽  
Parvaneh Azarali ◽  
Rostam Alizadeh
Keyword(s):  
Physical Activity ◽  
Environmental Factors ◽  
Sample Size ◽  
Dna Extraction ◽  
Gene Polymorphisms ◽  
Small Sample Size ◽  
Small Sample ◽  
Complex Phenotype ◽  
Genotype Frequencies ◽  
Pcr Rflp

Background: Physical activity is a complex phenotype influenced by millions of genes and environmental factors. It is well known that physical performance and sports ability are linked to genes variations. Objectives: The aim of this study was to evaluate the frequency of PGC1-α and CKMM gene polymorphisms in elite hockey athletes. Methods: Fifty Iranian elite national hockey athletes and 100 non-athletes participated in this study. To determine the genotypes of PGC1-α and CKMM gene polymorphisms, 2 ml of saliva was sampled and used for DNA extraction. To determine the genotypes, the PCR-RFLP method was employed. After examining the variants, the allele and genotype frequencies of subjects were measured. Results: The results showed no significant differences for the PPARGC1A gene in the percentage of AA, GG, and AG genotypes. Similarly, the percentage of these genotypes of the CKMM gene did not differ significantly between athletes and non-athletes. Conclusions: The results suggest that the gene polymorphisms of PGC1-α and CKMM are the same between the Iranian elite hockey athletes and non-athletes, which may be due to the overlapping effect of other genes and/or the small sample size of the study.

scholarly journals Identification of restriction enzyme in the FSHR gene of indonesian local cattle

2021 ◽  
Vol 888 (1) ◽  
pp. 012024
Author(s):  
P W Prihandini ◽  
A Primasari ◽  
M Luthfi ◽  
D Pamungkas ◽  
A P Z N L Sari ◽  
...  
Keyword(s):  
Restriction Enzyme ◽  
Restriction Site ◽  
Follicle Stimulating Hormone ◽  
Restriction Enzymes ◽  
Future Studies ◽  
Fshr Gene ◽  
Pcr Rflp ◽  
Mapping Analysis ◽  
Pcr Products ◽  
Enzyme Mapping

Abstract The restriction enzyme is important for genotyping using the PCR-RFLP technique. Therefore, this study aims to identify the restriction enzyme mapping in the partial sequence of the follicle-stimulating hormone receptor (FSHR) gene in Indonesian local cattle. A total of 29 samples sized 306 bp, were aligned with Genbank sequence acc no. NC_032660, resulting three polymorphic sites, namely g.193G>C, g.227T>C, and g.275A>C. Furthermore, the restriction mapping analysis using the NEBcutter program V2.0 showed that no enzyme recognized the SNP g.275A>C, while the SNP g.193G>C and g.227T>C were identified by the AluI and MscI enzymes, respectively. The AluI enzyme cuts at two positions (193 bp and 243 bp) in the G allele sample producing three fragments namely 50 bp, 63 bp, and 193 bp, meanwhile, in the C allele, the AluI cuts only in position 243 bp, hence, the fragment products are 63 bp and 243 bp. In contrast, the MscI enzyme was only recognized in the T allele, producing fragments sized 77 bp and 229 bp but failed to identify the restriction site along with the PCR products in the C allele. Based on the results, the SNPs (g.193G>C and g.227T>C) and restriction enzymes (AluI and MscI) are applicable for genotyping local Indonesian cattle using the PCR-RFLP technique in future studies.

Association between FVL G1691A, MTHFR C677T and A1298C Polymorphisms with Risk for Retinopathy of PrematurityAssociation between FVL G1691A, MTHFR C677T and A1298C Polymorphisms with Risk for Retinopathy of Prematurity

2021 ◽  
Author(s):  
Hamideh Shajari ◽  
Mohammadamin Ghadyani ◽  
Seyed Hamed Hosseini-Jangjou ◽  
Reza Bahrami ◽  
Seyed Alireza Dastgheib ◽  
...  
Keyword(s):  
Retinopathy Of Prematurity ◽  
Factor V Leiden ◽  
Mthfr C677t ◽  
Mthfr Gene ◽  
Factor V ◽  
Background Retinopathy ◽  
Increased Risk ◽  
Pcr Rflp ◽  
Rflp Assay ◽  
Preventable Blindness

Background: Retinopathy of prematurity (ROP) is an important cause of preventable blindness in children. The aim of this study was to examine the association of the polymorphisms at Factor V Leiden (FVL) and methylene tetrahydrofolate reductase (MTHFR) gene with risk of ROP. Methods: A total of 106 neonates with ROP and 110 healthy neonates were enrolled. The FVL G1691A and MTHFR C677T and A1298C polymorphisms were genotyped by PCR-RFLP assay. Results: There was a significant association between FVL G1691A polymorphism and an increased risk of ROP. However, the MTHFR C677T and A1298C polymorphisms were not associated with risk of ROP. Conclusion: FVL G1691A polymorphism may be risk factor for development of ROP in neonates. However, there was no significant association between MTHFR C677T and A1298C polymorphisms and risk of ROP. However, it is critical that larger and well-designed studies in different ethnicities are needed to confirm our conclusions.

scholarly journals C677T Polymorphism in the MTHFR Gene Is Associated With Risperidone-Induced Weight Gain in Schizophrenia

2020 ◽  
Vol 11 ◽  
Author(s):  
Jingping Liao ◽  
Ning Wang ◽  
Mengying Ma ◽  
Tianlan Lu ◽  
Hao Yan ◽  
...  
Keyword(s):  
Weight Gain ◽  
Mthfr Gene ◽  
C677t Polymorphism

scholarly journals Study of the Association between microRNA (miR-25T>C, miR-32C>A, miR-125C>T, and miR-222G>T) Polymorphisms and the Risk of Recurrent Pregnancy Loss in Korean Women

Genes ◽  
2020 ◽  
Vol 11 (4) ◽  
pp. 354 ◽  
Author(s):  
Jeong Yong Lee ◽  
Jung Oh Kim ◽  
Han Sung Park ◽  
Chang Soo Ryu ◽  
Ji Hyang Kim ◽  
...  
Keyword(s):  
Pregnancy Loss ◽  
Recurrent Pregnancy Loss ◽  
Rflp Analysis ◽  
Blood Coagulation Disorders ◽  
Single Nucleotide Variants ◽  
Genotype Combination ◽  
Nitrogen Levels ◽  
Genotype Frequencies ◽  
Pcr Rflp ◽  
Immunological Factors

Recurrent pregnancy loss (RPL), which is defined as two pregnancy losses that occur before 20 weeks of gestation, is relatively common, occurring in approximately 1–5% of women. The underlying cause is often unclear, although numerous factors may contribute to RPL, including environmental and immunological factors, blood coagulation disorders, and genetics. In particular, single nucleotide variants have been associated with RPL, including those found in microRNAs (miRNAs). We investigated the association between four miRNA polymorphisms, miR-25T>C, miR-32C>A, miR-125aC>T, and miR-222G>T, and RPL in a cohort consisting of 361 RPL patients and 272 controls. Subjects were genotyped at miRNA loci by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis, and genotype frequencies were calculated. We then performed allele and genotype combination analyses and measured the association between miRNA polymorphisms and clinical variables in both RPL patients and controls. We detected a statistically significant association between RPL and the miR-25T/miR-32C/miR-125aT/miR-222T allele combination (adjusted odds ratio (AOR), 4.361; 95% confidence interval (CI), 1.496–12.72; P = 0.003). Three-gene combinations, including miR-32C/miR-125aT/miR-222T (AOR, 3.085; 95% CI, 1.254–7.588; P = 0.010) and miR-25T/miR-125aT/miR-222T (AOR, 2.929; 95% CI, 1.183–7.257; P = 0.015), and the two-gene combination miR-125aT/miR-222T (AOR, 2.417; 95% CI, 1.084–5.386; P = 0.026) were also associated with RPL. Analysis of variance (ANOVA) revealed that platelet counts and blood urea nitrogen levels were significantly different in RPL patients expressing different miR-125aC>T and miR-25T>C genotypes, respectively (P < 0.05). In addition, creatinine levels were lower in RPL patients expressing the minor alleles miR-25T>C and miR-32C>A. We investigated miRNAs (miR-25, miR-32, miR-125a, miR-222) in RPL patients and healthy controls. Significantly different allele frequencies were detected by ANOVA. We suggest that miRNAs and clinical factors can impact RPL occurrence.

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