Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)

Abstract Background HLA‐G is a non‐classical class I gene of the human Major Histocompatibility encoding molecules with immune‐modulatory properties. Expression of HLA‐G is being largely studied in pathological conditions, such as tumors, viral infections, inflammation, and autoimmune diseases, graf...

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Autores principales:	Hamza Ben Salah, Refka Jelassi, Ines Zidi, Amor Ben Amor, Sondes Bizid, Radhia Ammi, Lamia Guizani, Aida Bouratbine, Karim Aoun, Hanen Chelbi
Formato:	article
Lenguaje:	EN
Publicado:	Wiley 2021
Materias:	Genetics QH426-470
Acceso en línea:	https://doaj.org/article/d44814da7182476d829379dd3d1db022
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spelling	oai:doaj.org-article:d44814da7182476d829379dd3d1db0222021-11-21T19:38:53ZRapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)2324-926910.1002/mgg3.1817https://doaj.org/article/d44814da7182476d829379dd3d1db0222021-11-01T00:00:00Zhttps://doi.org/10.1002/mgg3.1817https://doaj.org/toc/2324-9269Abstract Background HLA‐G is a non‐classical class I gene of the human Major Histocompatibility encoding molecules with immune‐modulatory properties. Expression of HLA‐G is being largely studied in pathological conditions, such as tumors, viral infections, inflammation, and autoimmune diseases, grafted tissues, among others. HLA‐G +3142C/G (rs1063320: dbSNP database) polymorphism is located in 3′ UTR of HAL‐G and plays a key role in determining the magnitude of gene and protein expression. The detection of HLA‐G +3142C/G polymorphism in the most published report is done through polymerase chain reaction followed by enzymatic digestion. Therefore, it is so interesting to develop a rapid and sensitive assay to genotype HLA‐G +3142C/G polymorphism. High‐resolution melt analysis (HRM) is a technology that is based on the analysis of the melting profile of PCR products through gradual temperature increase. The aim of this work is to apply high‐resolution melt method for genotyping the HLA‐G +3142C/G polymorphism. Methods DNA from 118 individuals was extracted from whole blood with QIAamp® DNA blood mini kit (Qiagen, Germany). Primer couple was designed using Primer 3 online tools so as to have only one SNP in the target sequence for high HRM efficiency. Positive Controls were identified using DNA sequencing and used as reference when assigning genotypes for trial samples. Results We were able to recognize the three genotypes with similar accuracy than DNA sequencing using high resolution melting method. Hardy‐Weinberg equilibrium test shows that our population is in equilibrium for the studied SNP. Genotypes frequencies of +3142C/G polymorphism in Tunisian general population are 0.475 for heterozygote G/C, 0.186 for homozygote G/G and 0.339 for homozygote C/C. Conclusion HRM is a cost‐effective method suitable for SNP genotyping.Hamza Ben SalahRefka JelassiInes ZidiAmor Ben AmorSondes BizidRadhia AmmiLamia GuizaniAida BouratbineKarim AounHanen ChelbiWileyarticleGeneticsQH426-470ENMolecular Genetics & Genomic Medicine, Vol 9, Iss 11, Pp n/a-n/a (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Genetics QH426-470
spellingShingle	Genetics QH426-470 Hamza Ben Salah Refka Jelassi Ines Zidi Amor Ben Amor Sondes Bizid Radhia Ammi Lamia Guizani Aida Bouratbine Karim Aoun Hanen Chelbi Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)
description	Abstract Background HLA‐G is a non‐classical class I gene of the human Major Histocompatibility encoding molecules with immune‐modulatory properties. Expression of HLA‐G is being largely studied in pathological conditions, such as tumors, viral infections, inflammation, and autoimmune diseases, grafted tissues, among others. HLA‐G +3142C/G (rs1063320: dbSNP database) polymorphism is located in 3′ UTR of HAL‐G and plays a key role in determining the magnitude of gene and protein expression. The detection of HLA‐G +3142C/G polymorphism in the most published report is done through polymerase chain reaction followed by enzymatic digestion. Therefore, it is so interesting to develop a rapid and sensitive assay to genotype HLA‐G +3142C/G polymorphism. High‐resolution melt analysis (HRM) is a technology that is based on the analysis of the melting profile of PCR products through gradual temperature increase. The aim of this work is to apply high‐resolution melt method for genotyping the HLA‐G +3142C/G polymorphism. Methods DNA from 118 individuals was extracted from whole blood with QIAamp® DNA blood mini kit (Qiagen, Germany). Primer couple was designed using Primer 3 online tools so as to have only one SNP in the target sequence for high HRM efficiency. Positive Controls were identified using DNA sequencing and used as reference when assigning genotypes for trial samples. Results We were able to recognize the three genotypes with similar accuracy than DNA sequencing using high resolution melting method. Hardy‐Weinberg equilibrium test shows that our population is in equilibrium for the studied SNP. Genotypes frequencies of +3142C/G polymorphism in Tunisian general population are 0.475 for heterozygote G/C, 0.186 for homozygote G/G and 0.339 for homozygote C/C. Conclusion HRM is a cost‐effective method suitable for SNP genotyping.
format	article
author	Hamza Ben Salah Refka Jelassi Ines Zidi Amor Ben Amor Sondes Bizid Radhia Ammi Lamia Guizani Aida Bouratbine Karim Aoun Hanen Chelbi
author_facet	Hamza Ben Salah Refka Jelassi Ines Zidi Amor Ben Amor Sondes Bizid Radhia Ammi Lamia Guizani Aida Bouratbine Karim Aoun Hanen Chelbi
author_sort	Hamza Ben Salah
title	Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)
title_short	Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)
title_full	Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)
title_fullStr	Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)
title_full_unstemmed	Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)
title_sort	rapid high‐resolution melting method to identify human leukocyte antigen‐g (hla‐g) 3′ untranslated region polymorphism +3142c/g (rs1063320)
publisher	Wiley
publishDate	2021
url	https://doaj.org/article/d44814da7182476d829379dd3d1db022
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Rapid high‐resolution melting method to identify human leukocyte antigen‐G (HLA‐G) 3′ untranslated region polymorphism +3142C/G (rs1063320)

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