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Identifier

000452159

Title

Detection of Sickle Cell Disease point-mutation for application at the point-of -care

Alternative Title

Ανίχνευση της σημειακής μετάλλαξης της δρεπανοκυτταρικής αναιμίας για εφαρμογή στο χώρο παροχής φροντίδας

Author

Δημητριάδη, Άννα Ε.

Thesis advisor

Γκιζελή, Ηλέκτρα

Reviewer

Στρουμπούλης, Ιωάννης
Καρδάσης, Δημήτρης

Abstract

Haemoglobinopathies refer to a diverse group of inherited disorders characterized by a reduced synthesis of one or more globin chains (thalassemia) or the synthesis of a structurally abnormal hemoglobin (Hb). Sickle cell disease (SCD) is an autosomal recessive haemoglobinopathy resulting from a single nucleotide polymorphism (SNP) leading to one nucleotide substitution (GTG for GAG) in the 6th codon of the bglobin gene. In sub-Saharan Africa, the Hb disorders are mainly associated to the pathological variants hemoglobin S (HbS) and HbC. Most applicable diagnostic techniques are based on amino acid detection and often require advanced laboratory equipment. Therefore, it is challenging to acquire a DNA-based and affordable diagnosis of SCD in sub-Saharan Africa. The aim of this study was to develop a molecular assay that targets the point-mutation within HbS gene. Two molecular assays were proposed; one primer-specific LAMP assay and one Restriction Enzyme-RPA assay (RE-RPA). Both molecular reactions are isothermal and widely used at the point-of-care. The specific LAMP assay was combined with a novel portable device that allows real-time colorimetric LAMP experiments. The RE-RPA assay was combined with the acoustic technology of Quartz Crystal Microbalance with Dissipation. The first assay enabled the discrimination of the wild type genotype (HbAA) and with the second assay the HbSS mutant genotype differentiation from the HbAA was achieved. By furtherly optimizing the RE-RPA assay, we hope to subsequently demonstrate a one-step efficient DNA amplification with high reliability at the point-of-care.

Language

English

Subject

Biosensor

Isothermal reaction

Βιοαισθητήρας

Ισοθερμική αντίδραση

Issue date

2022-11-25

Collection

School/Department--School of Sciences and Engineering--Department of Biology--Post-graduate theses

Type of Work--Post-graduate theses