Molecular Genetic Analysis of the Autosomal Recessive Non-Syndromic Inherited Retinitis Pigmentosa

Faiza Habib; Muhammad Yasin; Namal; Shaheryar; Areej Nasir; Abrar Hussain; Chinonso Ndubuisi; Hiba Azam; Muhammad Sajid; Arsalan Rasheed

doi:10.7759/cureus.37933

Molecular Genetic Analysis of the Autosomal Recessive Non-Syndromic Inherited Retinitis Pigmentosa

Cureus. 2023 Apr 21;15(4):e37933. doi: 10.7759/cureus.37933. eCollection 2023 Apr.

Authors

Faiza Habib¹, Muhammad Yasin², Namal³, Shaheryar⁴, Areej Nasir⁵, Abrar Hussain⁶, Chinonso Ndubuisi⁷, Hiba Azam⁸, Muhammad Sajid⁹, Arsalan Rasheed¹⁰

Affiliations

¹ Department of Molecular Biology and Genetics, Institute of Basic Medical Sciences, Khyber Medical University, Peshawar, PAK.
² Department of Biotechnology and Genetic Engineering, Kohat University of Science and Technology, Kohat, PAK.
³ Internal Medicine, United Medical and Dental College, Faisalabad, PAK.
⁴ Medicine, University College of Medicine and Dentistry, Lahore, PAK.
⁵ Medicine, Quaid-e-Azam Medical College, Bahawalpur, PAK.
⁶ Department of Biological Sciences, International Islamic University, Islamabad, PAK.
⁷ Family Medicine, Humboldt Park Health, Chicago, USA.
⁸ Surgery, University College of Medicine and Dentistry, Lahore, PAK.
⁹ Department of Biotechnology and Genetic Engineering, International Islamic University, Islamabad, PAK.
¹⁰ Department of Zoology, Faculty of Life and Chemical Sciences, Abdul Wali Khan University Mardan, Mardan, PAK.

Abstract

Introduction: 90% of visually impaired people live in developing countries. There are various types of vision impairment, but the focus of the current study is retinitis pigmentosa (RP). Up to now, 150 mutations have been reported that are linked with RP.

Methodology: Healthy and affected members from two Pakistani families (RP01 and RP02) segregating autosomal recessive RP were selected for DNA extraction. PCR was conducted, and the amplified PCR products were analyzed using Polyacrylamide Gel Electrophoresis (PAGE) and visualized in the Gel Doc system for linkage analysis. The Gene Hunter 2.1r5 tool in the Simple Linkage v5.052 beta software suite was used to conduct multipoint parametric linkage analysis on the two consanguineous families examined on the 6K Illumina array. Exons and intron-exon borders of all known arRP genes found in homozygous areas were sequenced in the matching probands using a 3130 automated sequencer and the Big Dye Terminator Cycle Sequencing Kit v3.1. The mutation study was carried out using the AlaMut 1.5 program.

Results: In both families, linkage analysis was performed using microsatellite marker DIS422 for gene crumbs homolog 1 (CRB1) and microsatellite marker D8S2332 for gene Retinitis Pigmentosa 1 (RP1). Multipoint linkage analysis identifies genomic regions that could potentially contain the genetic defect. In family RP01, only a single peak with a maximal multipoint LOD score of 3.00 was identified on chromosome 1, whereas in family RP02, multiple peaks with multipoint LOD scores of 1.80 were identified on chromosome 8. Analysis of the CRB1 gene revealed a homozygous substitution of glycine for valine (c.1152T>G; p.V243G), whereas the RP1 gene demonstrated that leucine was substituted for proline as a result of cytosine to thymine transfer (c.3419C>T; p. P1035L). Conclusion: Homozygosity mapping is a powerful method for finding genetic abnormalities that are both precise and comprehensive for identifying harmful variations in consanguineous families. This method is invaluable for providing accurate clinical diagnostic and genetic advice in remote regions of Pakistan while also increasing knowledge about autosomal recessive diseases and the dangers of mixing.

Keywords: crumbs homolog 1 (crb1); genetic mutation; linkage analysis; pakistan; retinitis pigmentosa; retinitis pigmentosa 1 (rp1); visual impairment.