An artificial-restriction fragment length polymorphism (A-RFLP) method for genotyping intronic SNP rs7587633 C/T of ATG16L1

Abstract

SNPs could either cause a disorder or directly alter the efficacy of a particular treatment and act as biological markers. The SNP rs7587633 C/T present in the intronic region of the ATG16L1 gene has been studied for its role in psoriasis vulgaris and Palmoplantar pustulosis. To genotype rs7587633 C/T using PCR-RFLP no restriction site is present for any of the restriction enzymes at the SNP position. To develop an artificial-RFLP method for genotyping rs7587633 C/T, the forward primer was designed in such a way that it resulted in the creation of an EcoRI restriction site in the amplified product which could further be digested with EcoRI to find the genotype of the individual. The newly developed A-RFLP method was applied to genotype the SNP rs7587633 C/T in DNA samples of 100 healthy control individuals. The allelic and genotypic frequencies of the SNPs were 0.80(C), 0.20(T) and 65%(CC), 31%(CT) and 4%(TT), respectively. In conclusion, we developed an A-RFLP method to genotype the SNP rs7587633 C/T which is not present in any of the natural restriction sites and this method could be applied to genotype this SNP in various populations/diseases to find its role.

Keywords:

• A-RFLP
• Intronic SNP
• rs7587633
• ATG16L1

Acknowledgement and declaration

The facilities and resources provided by Jaypee University of Information Technology, Solan, Himachal Pradesh, India to carry out this research are duly acknowledged. The help in the form of blood samples from the participants is also acknowledged.

Ethical approval

The study was approved by The Institutional Ethical Committee (IEC/project no.10-2014) of the Jaypee University of Information Technology, Solan, Himachal Pradesh, India approved this study.

Consent to participate

All the study subjects were provided with study-related information and enrolled after they provided written informed consent for their participation.

Author’s contribution

Ambika Sharma: performed and analyzed experimental data, writing the first draft of the manuscript. Arpna: analyzed experimental data, writing the first draft. Arti Sharma: writing the first draft of the manuscript. Jyoti Parkash: writing the first draft of the manuscript. Harish Changotra: Study concept and design, analyzed experimental data, edited the first draft of the manuscript, and supervised.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

Data available within the article or its supplementary materials.

Additional information

Funding

HC received funding from the Department of Science and Technology (SB/FT/LS-440/2012), Government of India for this research work.