Single nucleotide polymorphism detection by polymerase chain reaction-restriction fragment length polymorphism

Ôta, Masao; Fukushima, Hirofumi; Kulski, Jerzy K.; Inoko, Hidetoshi

doi:10.1038/nprot.2007.407

Cited by 107 publications

(75 citation statements)

References 46 publications

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“…An MboI recognition site was created by the MTNR1C SNP. The PCR products of this locus were digested with the restriction enzyme MboI (BBI Enzymes (UK) Ltd., Pontypool, UK) at 37°C for 16 h, ran on a 2% agarose gel, and stained with ethidium bromide (Ota et al, 2007) to detect the SNP by cleavage of the MTNR1C amplicon. All PCR products were purified with an EZ Spin Column DNA Gel Extraction Kit (Shanghai Sangon Biological Engineering Technology & Services Co. Ltd., Shanghai, P.R.…”

Section: Snp Discovery and Genotypingmentioning

confidence: 99%

Genetic effects of melatonin receptor genes on chicken reproductive traits

Li¹,

Zhang²,

Smith³

et al. 2013

Czech J. Anim. Sci.

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ABSTRACT:The melatonin receptors are G protein-coupled receptors (GPCR) that bind melatonin. Three types of melatonin receptors have been cloned. The MTNR1A (or Mel1A or MT1) and MTNR1B (or Mel1B or MT2) receptor subtypes are present in humans and other mammals, while an additional melatonin receptor subtype MTNR1C (or Mel1C or MT3) has been identified in amphibians and birds. Previous research has shown that the three common melatonin receptors regulate physiological processes, including seasonal reproduction and ovarian physiology. However, whether or not any polymorphisms of the different melatonin receptor subtypes are associated with reproductive traits in chickens is not known. In this study, we performed candidate gene analysis to identify single-nucleotide polymorphisms (SNPs) in the MTNR1A, MTNR1B, and MTNR1C genes in the Erlang Mountain Chicken population. SNP discovery was achieved by sequencing pooled DNA samples. Direct PCR-sequencing, PCR-SSCP/PCR-sequencing, and PCR-RFLP method were used to genotype the MTNR1A, MTNR1B, and MTNR1C genes, respectively. The GLM Procedure was used to estimate the statistical significance of association between genotypes at each locus and reproductive traits of chickens. In a sample of 460 chickens, four novel polymorphisms (JQ249890:g.384T>C, JQ249891:g.387T>C, JQ249894:g.63C>T, and JQ249896:g.294G>A) were detected in the melatonin receptor genes MTNR1A, MTNR1B, and MTNR1C, respectively. A statistically significant association (P < 0.01) was found between two SNPs (MTNR1A SNP, MTNR1C SNP) and reproductive traits: egg number at 300 days of age (EN) and age at first egg (AFE).

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Section: Snp Discovery and Genotypingmentioning

confidence: 99%

Genetic effects of melatonin receptor genes on chicken reproductive traits

Li¹,

Zhang²,

Smith³

et al. 2013

Czech J. Anim. Sci.

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“…Sometimes RFLP fragments are not observed as expected, so the unexpected outcome might be due to many different reasons, such as, if digestion of the PCR products is inefficient, the efficiency of the restriction enzyme reaction is poor or inactive, and some restriction enzymes are inhibited by methylation of nucleotides within their recognition sequences (Roberts et al, 2007) and the activity of some restriction enzymes (Ota et al, 2007). In spite of these problems that concern us about the sensitivity of the RFLP assay, unexpected results were not found when it was adjusted to 41.4 nM of plasmid DNA and no more than 6 mL of PCR product in the RFLP assay on RFLP.…”

Section: Fig 2 Tlr4mentioning

confidence: 90%

“…Restriction fragment length polymorphism (RFLP) is a classic method for SNP genotyping. It involves DNA amplification by polymerase chain reaction (PCR) and cleavage of the products by specific endonucleases generating a pattern of fragments that is easily identified by gel electrophoresis (Ota et al, 2007). Some SNPs are located in regions that are not recognized by known endonucleases.…”

Section: Introductionmentioning

confidence: 99%

Detection of theTLR41196C>T Polymorphism by Mismatched-Polymerase Chain Reaction Using Plasmid DNA as Internal Control in Restriction Fragment Length Polymorphism Assays

Lima-Neto

Hirata

Luchessi

et al. 2009

Genetic Testing and Molecular Biomarkers

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Blood samples were collected from 102 individuals with acute myocardial infarction (AMI) and 108 non-AMI individuals (controls) for DNA extraction and laboratory analyses. The 1196C > T polymorphism in the toll-like receptor 4 (TLR4) gene was amplified by mismatched-polymerase chain reaction (PCR). Amplicons and pBluescript II SK- plasmid were simultaneously digested with endonuclease HincII. Fragments were separated on 2% agarose gels. Plasmid was completely digested using up to 55.2 nmL/L DNA solutions and 1 microL PCR product. Nevertheless, plasmid DNA with 41.4 nM or higher concentrations was incompletely digested in the presence of 7 microL PCR product. In standardized conditions, TLR4 1196C>T variant was accurately genotyped. TLR4 1196T allele frequency was similar between AMI (3.1%) and controls (2.0%, p = 0.948). TLR4 SNP was not associated with AMI in this sample population. In conclusion, the plasmid-based control is a useful approach to prevent mistyping in RFLP assays, and it is validate for genetic association studies such as TLR4 1196C>T.

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“…Agarose gel (0.7%) was also employed to determine the quality and quantity of the extracted DNA, and the SNPs were identified through the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) technique [11]. The exon 2 segment related to the FokI, and the intron 8 segment targeted by BsmI were proliferated by the PCR, using special primers (Table 1).…”

Section: Dna Extractionmentioning

confidence: 99%

FokI and BsmI Polymorphisms of the VDR Gene and Breast Cancer Risk

Talaneh¹,

Ghorbani²,

Bakhshaiesh³

et al. 2016

mcij

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Introduction: Vitamin D fulfills its crucial role in cell proliferation and death through signal transduction into the nucleus by vitamin D receptor (VDR). Recent studies have depicted the association between VDR gene polymorphisms and different cancers, including breast cancer. This study attempted to consider the relationship between VDR gene polymorphisms and breast cancer risk among women in West Azerbaijan Province, Iran. Methods: FokI and BsmI polymorphisms were studied through the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique on blood samples taken from 95 cancer and 71 healthy participants from Urmia City. The association between the mentioned polymorphisms and breast cancer risk was considered using calculated odds ratio (OR). Results: The findings showed the protective effect of the FokI polymorphism in breast cancer development (OR = 0.2731), while no statistically significant correlation was observed between breast cancer and BsmI polymorphism (P > 0.05). Conclusions: This study identified the protective effect of the FokI polymorphism in breast cancer, and the absence of statistically significant association between BsmI polymorphism and breast cancer.

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Single nucleotide polymorphism detection by polymerase chain reaction-restriction fragment length polymorphism

Cited by 107 publications

References 46 publications

Genetic effects of melatonin receptor genes on chicken reproductive traits

Genetic effects of melatonin receptor genes on chicken reproductive traits

Detection of theTLR41196C>T Polymorphism by Mismatched-Polymerase Chain Reaction Using Plasmid DNA as Internal Control in Restriction Fragment Length Polymorphism Assays

FokI and BsmI Polymorphisms of the VDR Gene and Breast Cancer Risk

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