Alavian Seyed Ehsan, Sharafi Heidar, Shirmast Paniz, Alavian Seyed Moayed, Behnava Bita, Pouryasin Mohammad, Keshvari Maryam, Pouryasin Ali
Baqiyatallah Research Center for Gastroenterology and Liver Diseases, Baqiyatallah University of Medical Sciences, Tehran, Iran.
Middle East Liver Diseases (MELD) Virology Laboratory, MELD Center, Tehran, Iran.
J Clin Lab Anal. 2018 Sep;32(7):e22440. doi: 10.1002/jcla.22440. Epub 2018 Apr 16.
Inosine triphosphate pyrophosphatase (ITPA) gene single nucleotide polymorphisms (SNPs), rs1127354 and rs7270101, may cause a functional impairment in ITPase enzyme, resulting anemia protection in patients with chronic hepatitis C virus (HCV) infection undergoing ribavirin (RBV)-dependent regimens. The main purpose of this study was to provide and validate a simple, rapid, and inexpensive polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique for genotyping of ITPA rs1127354 and rs7270101 polymorphisms in chronic HCV-infected patients.
In the current study, 100 Iranian patients with chronic hepatitis C were examined and genotyped for ITPA rs1127354 and rs7270101 gene polymorphisms. To genotype rs1127354 and rs7270101 polymorphisms, PCR-RFLP technique and sequencing technique were performed on these samples. To validate the PCR-RFLP method, the PCR-RFLP genotyping results should be 100% concordant with the PCR-sequencing results.
The rs1127354 and rs7270101 polymorphisms of ITPA gene were genotyped by PCR-RFLP technique and sequencing simultaneously, and the results of both techniques were 100% concordant in all 100 patients. Both PCR-RFLP and sequencing techniques indicated that the genotypic frequency of rs7270101 was 80% AA, 19% AC and 1% CC, and for rs1127354 was 79% CC, 20% CA and 1% AA, respectively.
We developed and validated a rapid and inexpensive PCR-RFLP technique for the detection of ITPA rs1127354 and rs7270101 gene polymorphisms.
三磷酸肌苷焦磷酸酶（ITPA）基因单核苷酸多态性（SNP），即rs1127354和rs7270101，可能导致ITP酶功能受损，从而使接受基于利巴韦林（RBV）方案治疗的慢性丙型肝炎病毒（HCV）感染患者获得贫血保护作用。本研究的主要目的是提供并验证一种简单、快速且廉价的聚合酶链反应-限制性片段长度多态性（PCR-RFLP）技术，用于对慢性HCV感染患者的ITPA rs1127354和rs7270101多态性进行基因分型。
在本研究中，对100例伊朗慢性丙型肝炎患者进行检查，并对ITPA rs1127354和rs7270101基因多态性进行基因分型。为了对rs1127354和rs7270101多态性进行基因分型，对这些样本进行了PCR-RFLP技术和测序技术检测。为验证PCR-RFLP方法，PCR-RFLP基因分型结果应与PCR测序结果100%一致。
通过PCR-RFLP技术和测序同时对ITPA基因的rs1127354和rs7270101多态性进行基因分型，两种技术的结果在所有100例患者中均100%一致。PCR-RFLP和测序技术均表明，rs7270101的基因型频率分别为80%AA、19%AC和1%CC，rs1127354的基因型频率分别为79%CC、20%CA和1%AA。
我们开发并验证了一种快速且廉价的PCR-RFLP技术，用于检测ITPA rs1127354和rs7270101基因多态性。
