People who are working with Next Generation Sequencing for more than a decade now also get amused by its endless applications. The majority of researchers feel that only 20-25% of the total potential of Next Generation Sequencing has only been identified till now and a lot of areas are yet to be explored.
The Next Generation Sequencing is a more successful technology when we are using it for the analysis of the genomes of Humans or any other organism for that matter. It is not the only Genome Sequencing Technology before the introduction of Next Generation Sequencing there used to be another technology known as Sanger Sequencing. Sanger Sequencing was a very powerful tool for genome analysis It was highly effective, best for short reads, its accuracy was 99.99%, and so on.
But if we talk about the drawbacks of Sanger Sequencing is that it was expensive, the number of sequencing runs was also limited, and it was very tough to run it for sequences that do not have reference genomes. Hence, the introduction of Next Generation Sequencing enabled us to run the sequencing for a greater number of runs, we can do the sequencing of the genomes even if there is no reference genome and a lot of other features make Next Generation Sequencing the technology of future.
Till now we have talked about the technology, now let’s see how it works. But before that let’s see how many types of Next Generation Sequencing technologies are.
Applications of Next Generation Sequencing Technology
Whole Genome Sequencing
This analysis is run on a DNA Sample. In this technology, we are taking the genomic samples and running the analysis for both codings as well as non-coding regions i.e. Introns & Exons. This technology is found to be very effective in the case of Bacterial genomes where we can run the analysis to drug resistance, virulence & novel drug target, etc. It is one of the most powerful tools when it comes to genomic analysis as in this case are analyzing the whole genome without leaving any portion of it.
Transcriptome Analysis
This analysis is run on an RNA Sample. In this technology, we are analyzing the data and running the analysis to find differentially expressed genes in samples of our interest i.e. we are looking for unique genes expressed in either of the chosen samples. An example of its utility is that the COVID vaccines we have got are the RNA vaccines.
T-Cell Repertoire Sequencing
This analysis is run on an RNA Immunome Sample Data. With this technology, we can get a lot of information about the sample data. This technology has shown a lot of potential in detecting cancers and mutations related to different cancer types. We can use this technology in designing different vaccines also, after finding the mutations in the sample we can use the information for designing novel vaccine candidates.
Metagenomics Analysis
This analysis is run on DNA/RNA samples of Micro-Organisms. This technology specializes in the analysis of genomic information about the microbes. Like Soil Microbes, microbes in an environment like air, water Etc. It can help us in the analysis of soil samples from the Moon or even other planets also and help us in carrying out a lot of different analyses.
We have discussed Next Generation Sequencing and its different applications. Now let's look into the brief of the part of the analysis as to how we carry out the analysis of the pipeline used for the analysis.
Whenever we are carrying out the process of Next Generation Sequencing only 30% of the total work is carried out in the wet lab whereas the majority (70%) of work is carried out in the dry lab. To understand the process included in carrying out the wet lab process you can watch the informative animation of the process HERE. Now, let's talk about 70% of the work i.e. carrying out the Analysis of the sample with the help of Bioinformatics tools.
Analyzing the Next Generation Sequencing Samples
For different types of samples, there are different types of pipelines utilized when it comes to analyzing the genomic samples, but the initial process remains the same. So first let’s look at the complete overview of the process before we get into separate/different types of analysis.
This is the complete overview of the Next Generation Sequencing Data Analysis Pipeline. Where first and foremost step is to analyze the quality of the data received from the sequencer. Then in the very next step, we need to perform trimming of the data to improve the quality of the sequence. The low quality of the sequence is due to the presence of bad quality reads, added adapters (during sample preparation in the wet lab) Etc. which are the indicators of bad quality in the samples.
Hence in the trimming process, we remove all of these from our sample to improve the quality of our sample as now our sequence only contains the read related to our sample only. But again, after the trimming process, we check the quality of our sequences to check the difference in the quality of the sample before and after the trimming process.
Proceeding further next we perform the Mapping of our sequences with the reference genome which is the most important step as this step onward we start narrowing down to reach our desired results. So once the mapping is completed, we proceed with removing the unmapped reads.
Then after this step, we diversify by using different tools or approaches based on our sample types. For example, if we are working with DNA Samples so we will start with the Variant Analysis, in case we are working with RNA Samples so we will start with Transcriptome Analysis (for finding Differentially expressed genes) Etc.
So, the main motive behind this article is to make people, especially students aware of the huge potential next Generation Sequencing Carries and how one can perform the process. To know more about the Genome Analysis you can read it HERE.
Also, if you want to better understand the process included in the analysis of the sequencing data you can join us for a 3 Hour Short Course on Genome Analysis, you can register HERE
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