Particularly interesting is the single-molecule sequencing, which ensures that time and money are saved by skipping the initial cloning or amplification steps in NGS DNA sequencing. Nowadays, single-molecule DNA sequencing (also called third generation sequencing) is in the propagation phase and there is a limited number of research publications describing its use and applications. Since NGS DNA sequencing has many research applications, manufacturers did not give much importance to how this technology can be used in the clinic. In 2010, however, new platforms for clinical diagnostic laboratories have emerged. DNA microarrays (DNA chips) are 2B grids in which the nucleic acids in certain order are sequenced in a high density. Each point (from ~ 102 to 106 points in any sequence) represents a DNA probe that is attached to an inert surface of the glass or silicone. High-throughput screening of gene expression may reveal molecular signatures of events occurring at the cellular level. This information can be used in the clinic for diagnostic purposes or in research to understand the onset and progression of diseases. In microarrays, at least a two-fold increase in increased gene expression is considered as a threshold value of at least 0.5 times for decreased gene expression. Gene expression studies in microarray chips are considered only as a pre-study or screening. The results obtained must be verified with more specific measurements, such as real-time quantitative PCR The gene expression level microarray chips, which allow the transcriptome (all RNA species in a particular cell) to be examined and compared with the transcriptome in another cell, have proven successful in both research and clinical service. In this type of analysis it is possible to measure the amount of any desired mRNA.
Differences in expression may help explain tumor biology or identify tumor-specific targets for better diagnosis or new drug development Commercially produced microarrays currently include a wide variety of genes (TP53, CYP450) or pathways (apoptosis) or organisms (E. coli gene sequence). The presence of more objective markers to guide treatment and prognosis will be invaluable for the diagnosis and treatment of many diseases, especially cancers. Clinical-based microarray testing approved by the FDA for breast cancer diagnosis MammaPrint® provides an example of what can be achieved in the clinic with today's microarray technology.
In clinical medicine, microarrays can be useful for:
1. Diagnostic verification and disease classification.
2. Selection of individualized treatment by analyzing the germline DNA of the individual and somatic cell DNA in the tumor tissue.
3. Better prognostic indicators discovered by analysis of tumor DNA.
All genome assays (GWAS) significantly increased the potential for detecting genetic markers or genes involved in complex genetic disorders.
Factors that make this possible:
Testing larger groups;
The need for less SNP by dividing the genome into haplotype blocks and thanks to microarrays, it is easier and cheaper to work with a large number of SNP at the same time.
Affymetrix SNP chips have been developed to ensure suitability in different communities. Since it is difficult to provide homogeneous hybridization for all SNP probes, the detection rate is optimized with the coverage provided by the replacement probes. As an alternative to this product, there are also microchannel chips in bead format, marketed under the name of Illuminains BeadChips. BeadChips can be supplied as a ready-made panel or personalized as needed. For example, there is a panel of more than 400 SNPs associated with cancer. Illumina allows both NGS DNA sequencing and SNP genotyping to be performed with the same device, providing flexibility in analytical platforms.
Assist. Prof. Dr. Necip Ozan TİRYAKİOĞLU
