Designing, Optimization and Validation of Tetra Primer ARMS-PCR Protocol for Genotyping Single Nucleotide Polymorphism rs4731702 (C/T) of KLF14 Gene Associated with Type 2 Diabetes Mellitus: A Study in San Luis, Argentina
New, quick, and inexpensive methods for genotyping KLF14 gene polymorphism through Tetra
Primer ARMS-PCR were developed in the present investigation. Single nucleotide
polymorphism (SNP) genotyping needs to be attempted to establish association
between the identified SNP with metabolic trait for identification of molecular
markers that can be used to identify individuals at risk of developing Diabetes
Mellitus Type 2. In the current study, we have successfully genotyped the SNP
rs4731702 (C/T) in KLF14 gene. Tetra
primer ARMS-PCR protocol was optimized and validated for this SNP with short
turn-around time and costs. The optimized techniques were tested on 60 samples
of controls and type 2 Diabetes Mellitus individuals. Samples with known
genotypes for the described gene, previously tested in duplicate using the sequencing
methods, were employed for validation of the assay. Upon validation, complete
concordance was observed between the Tetra Primer ARMS-PCR assays and the
sequencing results. These results highlight the ability of Tetra Primer
ARMS-PCR in genotyping of SNP in KLF14
gene. Our results provide direct evidence that Tetra Primer ARMS-PCR is a
rapid, reliable, and cost-effective method for SNP genotyping of KLF14 gene in type 2 Diabetes Mellitus
individuals.
Cite this paper
Alvarez, M. F. , Gomez, M. E. V. and Siewert, S. (2016). Designing, Optimization and Validation of Tetra Primer ARMS-PCR Protocol for Genotyping Single Nucleotide Polymorphism rs4731702 (C/T) of KLF14 Gene Associated with Type 2 Diabetes Mellitus: A Study in San Luis, Argentina. Open Access Library Journal, 3, e3270. doi: http://dx.doi.org/10.4236/oalib.1103270.
Sladek, R., et al. (2007) A Genome-Wide Association Study Identifies Novel Risk Loci for Type
2 Diabetes. Nature, 445, 881-885. https://doi.org/10.1038/nature05616
Zeggini,
E., et al. (2007) Replication of Genome-Wide
Association Signals in UK Samples Reveals Risk Loci for Type 2 Diabetes. Science,
316, 1336-1341. https://doi.org/10.1126/science.1142364
Saxena, R., et al. (2007) Genome-Wide Association
Analysis Identifies Loci for Type 2 Diabetes and Triglyceride Levels. Science,
316, 1331-1336. https://doi.org/10.1126/science.1142358
Steinthorsdottir, V., et al. (2007) A Variant in CDKAL1 Influences
Insulin Response and Risk of Type 2 Diabetes. Nature Genetics,
39, 770-775. https://doi.org/10.1038/ng2043
Zeggini, E., et al. (2008)
Meta-Analysis of Genome-Wide Association Data and Large-Scale Replication
Identifies Additional Susceptibility Loci for Type 2 Diabetes. Nature Genetics, 40,
638-645. https://doi.org/10.1038/ng.120
Yasuda, K., et al. (2008) Variants in KCNQ1 Are Associated with Susceptibility to Type 2 Diabetes
Mellitus.Nature Genetics, 40, 1092-1097. https://doi.org/10.1038/ng.207
Unoki, H., et al. (2008) SNPs in KCNQ1 Are Associated with Susceptibility to Type 2 Diabetes
in East Asian and European Populations. Nature
Genetics, 40, 1098-1102. https://doi.org/10.1038/ng.208
Rung, J., et al. (2009) Genetic Variant near IRS1 Is Associated with Type 2 Diabetes,
Insulin Resistance and Hyperinsulinemia. Nature
Genetics, 41, 1110-1115. https://doi.org/10.1038/ng.443
Kong, A., et al. (2009) Parental Origin of Sequence Variants Associated with Complex Diseases. Nature, 462, 868-874. https://doi.org/10.1038/nature08625
Dupuis,
J., et al. (2010) New Genetic Loci
Implicated in Fasting Glucose Homeostasis and Their Impact on Type 2 Diabetes
Risk. Nature Genetics, 42, 105-116. https://doi.org/10.1038/ng.520
Saxena, R., et al. (2010) Genetic Variation in GIPR Influences the Glucose and
Insulin Responses to an Oral Glucose Challenge. Nature Genetics, 42, 142-148. https://doi.org/10.1038/ng.521
Teslovich, T.M., et al. (2010) Biological, Clinical and Population Relevance of 95
Loci for Blood Lipids. Nature, 466, 707-713. https://doi.org/10.1038/nature09270
Chasman, D.I., et al. (2009) Forty-Three Loci Associated with Plasma Lipoprotein
Size, Concentration, and Cholesterol Content in Genome-Wide Analysis. PLoS Genetics, 5, e1000730. https://doi.org/10.1371/journal.pgen.1000730
Jeemon, P., et al. (2011) Implications of Discoveries from Genome-Wide
Association Studies in Current Cardiovascular Practice. World Journal of Cardiology, 3, 230-247. https://doi.org/10.4330/wjc.v3.i7.230
Ohshige, T., et al. (2011) Association of New Loci Identified in European
Genome-Wide Association Studies with Susceptibility to Type 2 Diabetes in the
Japanese. PLoS ONE, 6, e26911. https://doi.org/10.1371/journal.pone.0026911
Voight, B.F., et al. (2010) Twelve Type 2 Diabetes Susceptibility Loci Identified
through Large-Scale Association Analysis. Nature
Genetics, 42, 579-589. https://doi.org/10.1038/ng.609
Rees, S.D., et al. (2011) Replication of 13 Genome-Wide Associations
(GWA)-Validated Risk Variants for Type 2 Diabetes in Pakistani Populations. Diabetologia, 54, 1368-1374. https://doi.org/10.1007/s00125-011-2063-2
Small, K.S., et al. (2011) Identification of an imprinted Master Trans Regulator
at the KLF14 Locus Related to Multiple Metabolic Phenotypes. Nature Genetics, 43, 561-564. https://doi.org/10.1038/ng.833
Chen, X., et al. (2012) Genome-Wide Association Study Validation Identifies
Novel Loci for Atherosclerotic Cardiovascular Disease. Journal of Thrombosis and Haemostasis, 10, 1508-1514. https://doi.org/10.1111/j.1538-7836.2012.04815.x
Hanson, R.L., et al. (2013) Strong Parent-of-Origin Effects in the Association of
KCNQ1 Variants with Type 2 Diabetes in American Indians. Diabetes, 62, 2984-2991. https://doi.org/10.2337/db12-1767
Old, J.M., Varawalla, N.Y. and Weatherall, D.J.
(1990) Rapid Detection and Prenatal Diagnosis of Beta-Thalassaemia: Studies in
Indian and Cypriot Populations in the UK. Lancet,
336, 834-837. https://doi.org/10.1016/0140-6736(90)92338-I
Vannucchi, A.M., Pancrazzi, A, Bogani, C, Antonioli, E and Guglielmelli, P. (2006)
A Quantitative Assay for JAK2V617F Mutation in Myeloproliferative Disorders by ARMS-PCR and Capillary Electrophoresis. Leukemia, 20, 1055-1060. https://doi.org/10.1038/sj.leu.2404209
Wang, Y.Z., et al. (2014) Detection of Hepatitis B Virus A1762T/G1764A Mutant
by Amplification Refractory Mutation System. Brazilian Journal of Infectious Diseases, 18, 261-265. https://doi.org/10.1016/j.bjid.2013.09.005
Ye, S., Humphries, S. and Green, F. (1992) Allele Specific Amplification by Tetra-Primer
PCR. Nucleic Acids Research, 20, 1152. https://doi.org/10.1093/nar/20.5.1152
Black, A.R., Black, J.D. and Azizkhan-Clifford, J. (2001) Sp1 and Kruppel-Like Factor Family of Transcription
Factors in Cell Growth Regulation and Cancer. Journal of Cellular Physiology, 188, 143-160. https://doi.org/10.1002/jcp.1111
Sarmento,
O.F., et al. (2015) A Novel Role for
KLF14 in T Regulatory Cell Differentiation. Cellular
and Molecular Gastroenterology and Hepatology, 1, 188-202.e4. https://doi.org/10.1016/j.jcmgh.2014.12.007
De
Assuncao, T.M., et al. (2014) New
Role for Kruppel-Like Factor 14 as a Transcriptional Activator Involved in the
Generation of Signaling Lipids. Journal
of Biological Chemistry, 289,
15798-15809. https://doi.org/10.1074/jbc.M113.544346
Huang, P., et al. (2013) Association of the KLF14 rs4731702 SNP and Serum
Lipid Levels in the Guangxi Mulao and Han Populations. BioMed Research International, 2013, Article
ID: 231515. https://doi.org/10.1155/2013/231515
Chen, G., et al. (2012) Genome-Wide Association Study Identifies Novel Loci
Association with Fasting Insulin and Insulin Resistance in African Americans. Human Molecular Genetics, 21, 4530-4536. https://doi.org/10.1093/hmg/dds282
Stacey, S.N., et al. (2009) New Common Variants Affecting Susceptibility to Basal
Cell Carcinoma. Nature Genetics, 41, 909-914. https://doi.org/10.1038/ng.412
Steegenga, W.T., et al. (2014) Genome-Wide
Age-Related Changes in DNA Methylation and Gene Expression in Human PBMCs. Age, 36, 9648. https://doi.org/10.1007/s11357-014-9648-x
Zhao, Z. and Boerwinkle, E. (2002) Neighboring-Nucleotide Effects on Single Nucleotide Polymorphisms:
A Study of 2.6 Million Polymorphisms across the Human Genome. Genome Research, 12, 1679-1686. https://doi.org/10.1101/gr.287302
Etlik, O., Koksal, V., Arican-Baris, T.S.
and Baris, I. (2008) An Improved Tetra-Primer PCR Approach for the Detection of
the FGFR3 G380R Mutation Responsible for Achondroplasia. Molecular and Cellular Probes, 22, 71-75. https://doi.org/10.1016/j.mcp.2007.06.002
Siewert, S., Filipuzzi, S., Codazzi, L., Gonzalez, I.
and Ojeda, M. (2007) Impact of Metabolic Syndrome Risk Factors in First-Degree
Relatives of Type 2 Diabetic Patients. Review
of Diabetic Studies, 4, 177-184. https://doi.org/10.1900/RDS.2007.4.177