Sequencing is a crucial element of the process of discovering and studying genetics. It’s also changing rapidly, which is why we’ve created this list of useful information describing the various DNA & RNA sequencing and epigenomics.
Suppose you like this list of helpful DNA and RNA sequencing types. In that case, you may be interested in downloading the free Next Generation Sequencing Guide that includes this information, the latest background of NGS sequencing, the steps to sequencing, and more.
Types of DNA Sequencing
1. Whole Genome Sequencing (WGS)
Whole genome sequencing can be used to determine the sequence of the genome that an animal has. In addition, WGS can be used to identify variations (mutation) prevalent within populations of organisms and to link genetic variations with diseases through genome-wide association research (GWAS). A GWAS conducts WGS on two groups and then compares the differences in traits with genetic variations to link traits with known variants. WGS was first used for diagnostic purposes in 2009. However, time and cost have limited its application in this field. Since the cost of WGS has declined to increase, it’s becoming more popular as an instrument for diagnosing. After achieving a “$1000 genome “$1000 genome”, multiple companies are working toward the goal of achieving a “$100 genome” “$100 genome”.
2. Targeted sequencing
Targeted NGS lets users analyze specific parts of the genome to conduct in-depth analyses more quickly and cost-effectively than complete genome sequencing. Targeted sequencing can identify the existence of novel and well-known variants in the area of interest. It typically produces less information than WGS, which makes it easier to manage the analysis.
Many methods of targeted sequencing are suitable to the specific needs. The most well-known techniques are hybridization capture amplicon sequencing and molecular-inversion probes (MIPs). Check out the Targeted Sequencing Guide to learn more about the in-depth differences between hybridization capture and amplicon sequence.
3. Whole exome sequencing (WES)
Whole exome sequencing can identify all the protein-coding genes that are part of the genome. Concentrating on exons that code for proteins (and exclusion of other parts in the genome) could reduce the expense and time involved in sequencing because exons make up only one percent part of the genome. The variations in exons that encode proteins can be responsible for various diseases; therefore, this type of sequencing is usually enough for diagnostic purposes. WES is a better method of mapping the variants that are not common in the general population to help identify the causes of complex diseases. It’s also an excellent alternative for discovering research. WES is especially useful in research on cancer and is currently used for cancer diagnosis. The information gathered from WES can give insight into the future and personalized treatment options. WES is typically performed using hybridization probes rather than amplifiers.
4. Hybridization capture
Before hybridization capture, samples are transformed into libraries of sequencing. The regions of interest in the library are then retrieved with biotinylated oligonucleotide long-oligo. Since the DNA was randomly and sheared during library preparation, the fragments captured are unique and overlapped. Baits can be tiled, overlaid, and placed to address the challenges of repetitive sequences, etc. With the help of advanced technology, the capture process can be made extremely uniform. Hybridization capture is typically employed to target exome sequencing. Other applications include testing for genotyping, detection of rare variants, and oncology diagnostics.
5. Amplicon sequencing
Amplicon Sequencing is a targeted method that permits you to study the variation within particular genomic regions. This technique uses PCR to amplify DNA to create amplified samples. The amplicons are then indexed and sequenced. Amplicon sequencing is usually utilized for the detection of disease-associated variants and diagnostics. It is also utilized to genotype by sequencing and to confirm CRISPR genome editing. Find out more about assessing CRISPR-Cas9 changes quickly and precisely using rhAmpSeq-targeted DNA sequencing.
6. Probes for Molecular Inversion (MIPs)
Molecular inversion probes are yet another commonly used method of enriching target sequences. Specific sequences targeted for enrichment are joined at both ends, a universal sequence to create the MIP. The MIP forms a hybrid with the desired region before the gap-filling reaction occurs, and then a third ligation closes the circles. A restriction enzyme within the MIP could form a linear molecule. The sequences targeted are amplified before taking the sequence. MIPs are especially useful in large-scale genotyping.
Types of RNA Sequencing
- Whole transcriptome sequencing (WTS)
Because RNA expression levels will vary depending on the type of cell and the state of disease, Understanding the transcriptome could offer valuable information. Sequencing the entire transcriptome gives complete data, including information on noncoding and coding transcripts and known and new variations. One variant in Whole Transcriptome Sequencing, or RNA-seq, is called stranded, which preserves data about the strand from which the transcripts originate. Transcripts. This allows you to find new transcripts, such as antisense and antisense RNAs. The use of RNA-seq is typically for discovery research.
Typically, RNA-seq determines the messenger RNA (mRNA) expression level. It can also be used to determine changes in gene expression with time, such as gene fusions or mononucleotide polymorphisms (SNPs), alternative splicing, and RNA modifications.
2. Gene expression targeted by RNA-Sequencing
Like DNA, RNA could be detected for a sequence with hybridization capture and amplicon sequencing techniques. Certain groups of RNA could be targeted. It is usually the case with coding transcripts; however, other types of RNA, like the tRNA (transfer the RNA) and small RNA, might be enhanced.
3. Ribosomal RNA depletion
The ribosomal RNA (rRNA) is responsible for 80 to 95 percent of the cell’s total RNA. However, because its expression is consistent, this is not important. This is why it’s beneficial not to sequence these molecules and concentrate your sequence on more valuable information. The rRNA could remove itself from samples through two methods 1)). Biotinylated probes may bind to the rRNA and take it out of the sample. 2). DNA-based probes which bind to rRNA could be used in conjunction with RNase H to remove the rRNA from the library sample before library preparation.
Epigenomics
1. ChIP-Sequencing
Chromatin immunoprecipitation (ChIP) determines the protein’s interactions with DNA. Proteins can regulate DNA, changing its expression. The surrounding environment can affect this regulation and may alter in time. ChIP is a method of identifying the sites within the DNA sequence to which protein is attached. An antibody is employed to attach to the protein of importance, which allows the immunoprecipitation process of DNA bound with the protein.
If this identification is performed using an array, it’s called a ChIP chip. Evaluation of protein interactions across the entire genome through sequencing is known as ChIP-Seq. ChIP-Seq studies typically focus on studying transcription factors, histones, and histone modifications to provide insights into the regulation of genes, cell growth, and disease progression.
2. Methyl-seq
Methylation, a different method through which DNA expression is controlled, can also be affected by the environment and alter over time. Methyl groups are incorporated into DNA sequences and can inhibit DNA expression.
Methyl-Seq, also called bisulfite-sequencing, processes the DNA using bisulfite before sequencing to reveal the methylation status in your DNA. This information is utilized to determine the relationship between genes and environmental factors.
