Application of polymerase chain reaction-restriction fragment length polymorphism (RFLP-PCR) in the analysis of single nucleotide polymorphisms (SNPs)

Authors

DOI:

https://doi.org/10.18778/1730-2366.16.14

Keywords:

nucleotide polymorphisms, DNA analysis, polymerase chain reaction

Abstract

Polymerase chain reaction-restriction fragment length polymorphism (RFLP-PCR) is a technique used to identify single nucleotide polymorphisms (SNPs) based on the recognition of restriction sites by restriction enzymes. RFLP-PCR is an easy-to-perform and inexpensive tool for initial analysis of SNPs potentially associated with some monogenic diseases, as well as in genotyping, genetic mapping, lineage screening, forensics and ancient DNA analysis. The RFLP-PCR method employs four steps: (1) isolation of genetic material and PCR; (2) restriction digestion of amplicons; (3) electrophoresis of digested fragments; and (4) visualisation. Despite its obsolescence and the presence of high-throughput DNA analysis techniques, it is still applied in the analysis of SNPs associated with disease entities and in the analysis of genetic variation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). RFLP-PCR is a low-cost and low-throughput research method allowing for the analysis of SNPs in the absence of specialised equipment, and it is useful when there is a limited budget.

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References

Alavian, S.E., Sharafi, H., Shirmast, P., Alavian, S. M., Behnava, B., Pouryasin, M., Keshvari, M., Pouryasin, A. 2018. A facile PCR-RFLP method for genotyping of ITPA rs1127354 and rs7270101 polymorphisms. Journal of Clinical Laboratory Analysis, 32: e22440.
Google Scholar DOI: https://doi.org/10.1002/jcla.22440

Backfisch, W., Neuenschwander, S., Giger, U., Stranzinger, G., Pliška, V. 1994. Carrier detection of ovine hemophilia a using an RFLP marker, and mapping of the factor VIII gene on the ovine X-chromosome. Journal of Heredity, 85: 474–478.
Google Scholar DOI: https://doi.org/10.1093/oxfordjournals.jhered.a111503

Berg Rasmussen, H. 2012. Restriction fragment length polymorphism analysis of PCR-ampli-restriction fragment length polymorphism afied fragments (PCR-RFLP) and gel electrophoresis – valuable tool for genotyping and genetic fingerprinting. In: Magdeldin, S. (ed.) Gel Electrophoresis – Principles and Basics, InTech, London.
Google Scholar DOI: https://doi.org/10.5772/37724

Bhattacharyya, C., Das, C., Ghosh, A., Singh, A., Mukherjee, S., Majumder, P., Basu, A., Biswas, N. 2020. Global spread of SARS-CoV-2 subtype with spike protein mutation D614G is shaped by human genomic variations that regulate expression of TMPRSS2 and MX1 genes. bioRxiv, 2020.05.04.075911.
Google Scholar DOI: https://doi.org/10.1101/2020.05.04.075911

Budowle, B., Chakraborty, R., Giusti, A.M., Eisenberg, A.J., Allen, R.C. 1991. Analysis of the VNTR locus D1S80 by the PCR followed by high-resolution PAGE. American Journal of Human Genetics, 48: 137–144.
Google Scholar

Catamo, E., Zupin, L., Segat, L., Celsi, F., Crovella, S. 2015. HLA-G and susceptibility to develop celiac disease. Human Immunology, 76: 36–41.
Google Scholar DOI: https://doi.org/10.1016/j.humimm.2014.12.006

Chang, H.W., Cheng, Y.H., Chuang, L.Y., Yang, C.H. 2010. SNP-RFLPing 2: an updated and integrated PCR-RFLP tool for SNP genotyping. BMC Bioinformatics, 11: 173.
Google Scholar DOI: https://doi.org/10.1186/1471-2105-11-173

Chang, H.W., Yang, C.H., Chang, P.L., Cheng, Y.H., Chuang, L.Y. 2006. SNP-RFLPing: restriction enzyme mining for SNPs in genomes. BMC Genomics, 7: 30.
Google Scholar DOI: https://doi.org/10.1186/1471-2164-7-30

Endreffy, E., Várkonyi, Á., Kaiser, G.I., Raskó, I. 1992. Association of altered RFLP with coeliac disease among Hungarian families. Journal of Pediatric Gastroenterology and Nutrition, 14: 118–119.
Google Scholar DOI: https://doi.org/10.1097/00005176-199201000-00026

Hagelberg, E., Hofreiter, M., Keyser, C. 2015. Ancient DNA: the first three decades. Philosophical Transaction of the Royal Society B: Biological Sciences, 370: 20130371.
Google Scholar DOI: https://doi.org/10.1098/rstb.2013.0371

Harding, D. 2007. Impact of common genetic variation on neonatal disease and outcome. Archives of Disease in Childhood. Fetal and Neonatal Edition, 92: F408–F413.
Google Scholar DOI: https://doi.org/10.1136/adc.2006.108670

Hashemi, S.A., Khoshi, A., Ghasemzadeh-Moghaddam, H., Ghafouri, M., Taghavi, M., Namdar-Ahmadabad, H., Azimian, A. 2020. Development of a PCR-RFLP method for detection of D614G mutation in SARS-CoV-2. Infection, Genetics and Evolution, 86: 104625.
Google Scholar DOI: https://doi.org/10.1016/j.meegid.2020.104625

Herrmann, F.H., Wehnert, M., Wulff, K. 2008. RFLP analysis for diagnosis of haemophilia A in the German Democratic Republic. Clinical Genetics, 37: 12–17.
Google Scholar DOI: https://doi.org/10.1111/j.1399-0004.1990.tb03384.x

Kozák, L., Kuhrová, V., Blažková, M., Fajkusová, L., Dvořáková, D., Romano, V., Pijáčková, A. 1995. Phenylketonuria mutations and their relation to RFLP haplotypes at the PAH locus in Czech PKU families. Human Genetics, 96: 472–476.
Google Scholar DOI: https://doi.org/10.1007/BF00191809

Křepelová, A., Brdicka, R., Vorlová, Z. 1993. Factor VIII gene mutations and RFLP analysis in hemophilia A. Stem Cells, 11: 72–76.
Google Scholar DOI: https://doi.org/10.1002/stem.5530110615

Laber, T.L., Giese, S.A., Iverson, J.T., Liberty, J.A. 1994. Validation studies on the forensic analysis of restriction fragment length polymorphism (RFLP) on LE agarose gels without ethidium bromide: effects of contaminants, sunlight, and the electrophoresis of varying quantities of deoxyribonucleic acid (DNA). Journal of Forensic Sciences, 39: 13649J.
Google Scholar DOI: https://doi.org/10.1520/JFS13649J

Meijer, H., Jongbloed, R.J.E., Hekking, M., Spaapen, L.J.M., Geraedts, J.P.M. 1993. RFLP haplotyping and mutation analysis of the phenylalanine hydroxylase gene in Dutch phenylketonuria families. Human Genetics, 92: 588–592.
Google Scholar DOI: https://doi.org/10.1007/BF00420944

Orlando, L., Allaby, R., Skoglund, P., Sarkissian, C. Der, Stockhammer, P.W., Ávila-Arcos, M.C., Fu, Q., Krause, J., Willerslev, E., Stone, A.C., Warinner, C. 2021. Ancient DNA analysis. Nature Reviews Methods Primers 1: 1–26.
Google Scholar DOI: https://doi.org/10.1038/s43586-020-00011-0

Pingoud, A., Jeltsch, A. 2001. Structure and function of type II restriction endonucleases. Nucleic Acids Research, 29: 3705–3727.
Google Scholar DOI: https://doi.org/10.1093/nar/29.18.3705

Plante, J.A., Liu, Y., Liu, J., Xia, H., Johnson, B.A., Lokugamage, K.G., Zhang, X., Muruato, A.E., Zou, J., Fontes-Garfias, C.R., Mirchandani, D., Scharton, D., Bilello, J.P., Ku, Z., An, Z., Kalveram, B., Freiberg, A.N., Menachery, V.D., Xie, X., Plante, K.S., Weaver, S.C., Shi, P.Y. 2020. Spike mutation D614G alters SARS-CoV-2 fitness. Nature, 592: 116–121.
Google Scholar DOI: https://doi.org/10.1038/s41586-020-2895-3

Pramoonjago, P., Harahap, A., Taufani, R.A., Setianingsih, I., Marzuki, S. 1999. Rapid screening for the most common β thalassaemia mutations in south east Asia by PCR based restriction fragment length polymorphism analysis (PCR-RFLP). Journal of Medical Genetics, 36: 937–938.
Google Scholar

Saiki, R.K., Scharf, S., Faloona, F., Mullis, K.B., Horn, G.T., Erlich, H.A., Arnheim, N. 1985. Enzymatic amplification of β-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science, 230: 1350–1354.
Google Scholar DOI: https://doi.org/10.1126/science.2999980

Stankovic, M., Rakicevic, L., Mikovic, D., Jankovic, G., Nikolic, A. 2005. Indirect diagnosis of haemophilia B by multiplex PCR/RFLP. Clinical & Laboratory Haematoology, 27: 145–146.
Google Scholar DOI: https://doi.org/10.1111/j.1365-2257.2005.00671.x

Tasleem Raza, S., Husain, N., Kumar, A. 2009. Screening for hemophilia A carriers: Uutility of PCR-RFLP-based polymorphism analysis. Clinical and Applied Thrombosis, 15: 78–83.
Google Scholar DOI: https://doi.org/10.1177/1076029607305105

Zhou, B., Thi Nhu Thao, T., Hoffmann, D., Taddeo, A., Ebert, N., Labroussaa, F., Pohlmann, A., King, J., Steiner, S., Kelly, J.N., Portmann, J., Halwe, N.J., Ulrich, L., Trüeb, B.S., Fan, X., Hoffmann, B., Wang, L., Thomann, L., Lin, X., Stalder, H., Pozzi, B., Brot, S. de, Jiang, N., Cui, D., Hossain, J., Wilson, M., Keller, M., Stark, T.J., Barnes, J.R., Dijkman, R., Jores, J., Benarafa, C., Wentworth, D.E., Thiel, V., Beer, M. 2021. SARS-CoV-2 spike D614G change enhances replication and transmission. Nature, 592: 122–127.
Google Scholar DOI: https://doi.org/10.1038/s41586-021-03361-1

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Published

2021-09-29

How to Cite

Tarach, P. (2021). Application of polymerase chain reaction-restriction fragment length polymorphism (RFLP-PCR) in the analysis of single nucleotide polymorphisms (SNPs). Acta Universitatis Lodziensis. Folia Biologica Et Oecologica, 17, 48–53. https://doi.org/10.18778/1730-2366.16.14