Molecular genetic testing methodologies in hematopoietic diseases: current and future methods

Abstract

Introduction Rapid technological advancements in clinical molecular genetics have increased our diagnostic and prognostic capabilities in health care. Understanding these assays, as well as how they may change over time, is critical for pathologists, clinicians, and translational researchers alike. Methods This review provides a practical summary and basic reference for current molecular genetic technologies, as well as new testing methodologies that are in use, gaining momentum, or anticipated to contribute more broadly in the future. Results Here, we discuss DNA and RNA based methodologies including classic assays such as the polymerase chain reaction (PCR), Sanger sequencing, and microarrays, to more cutting‐edge next‐generation sequencing (NGS) based assays and emerging molecular technologies such as cell‐free DNA (cfDNA) or circulating tumor DNA (ctDNA), and NGS‐based detection of infectious disease organisms. Conclusion This review serves as a basic foundation for knowledge in current and emerging clinical molecular genetic technologies.

Cytogenetic and Biochemical Genetic Techniques for Personalized Drug Therapy in Europe

“…Especially in cancer treatment, the application of FISH as a diagnostic method to guide therapy with regard to genetic abnormalities is recommended or even determined in the summary of product characteristics for certain drugs as a confirmatory test method of choice if the IHC first-tier method results are equivocal ( Table 1 ). Thus, FISH assays are applied in clinical diagnostics and can be used to guide targeted therapy [ 19 , 20 ].…”

“…Thus, it is labor-intensive, time-consuming, technically demanding and relatively costly. However, analytically valid test kits for the analysis of different clinically relevant conditions are commercially available and thus can be applied directly after tissue preparation [ 19 ]. Chromogenic In Situ Hybridization (CISH) and Silver In Situ Hybridization (SISH) can also be used to examine the chromosomal number, chromosomal translocations and gene copy number variations due to gene amplification.…”

Genetic Markers in Lung Cancer Diagnosis: A Review

“… Polymerase chain reaction (PCR)—PCR allows for the exponential amplification of specific targeted genetic loci in a reaction mixture containing DNA primers, deoxynucleotides (dNTPs), and DNA polymerases. PCR is a qualitative technique to amplify and copy a targeted area of extracted DNA a million to a billion-fold over [ 49 ]. In the course of time, modifications and advances in this molecular diagnostics technique enabled the relative or even absolute quantification of DNA through the usage of quantitative real-time PCR (qPCR) or partition-based PCR techniques, such as droplet digital PCR (ddPCR) [ 27 , 49 ].…”

“…PCR is a qualitative technique to amplify and copy a targeted area of extracted DNA a million to a billion-fold over [ 49 ]. In the course of time, modifications and advances in this molecular diagnostics technique enabled the relative or even absolute quantification of DNA through the usage of quantitative real-time PCR (qPCR) or partition-based PCR techniques, such as droplet digital PCR (ddPCR) [ 27 , 49 ]. The exploitation of PCR in quantitative DNA analysis is leading to the increased clinical usefulness of PCR for a broad range of applications; PCR is used in a variety of methods, such as allele-specific PCR-based methods or mutation screening methods, including melting curve analysis, that are used in the analysis of mutations sequences.…”

“…FISH uses specific fluorescent probes that bind to nucleic acid sequences with a high degree of sequence complementarity, helping one to localize these DNA sequences on chromosomes. Microarray-based CGH (array CGH) enables us genome-wide screening for chromosomal imbalances on the basis of genomic DNA hybridization to complement probes that are immobilized on a slide [ 17 , 23 , 49 ].…”