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GT-FAR is an RNA seq pipeline that performs RNA-seq QC, alignment, reference free quantification, and splice variant calling. It filters, trims, and sequentially aligns reads to gene models and predicts and validates new splice junctions after which it quantifies expression for each gene, exon, and known/novel splice junction, and Variant Calling.
RNA-Seq (named as an abbreviation of RNA sequencing) is a technique that uses next-generation sequencing to reveal the presence and quantity of RNA molecules in a biological sample, providing a snapshot of gene expression in the sample, also known as transcriptome.
The MIQE guidelines were created due to the low quality of qPCR data submitted to academic journals at the time, which was only becoming more common as Next Generation Sequencing machinery allowed for such experiments to be run for a cheaper cost. Because the technique is utilized across all of science in multiple fields, the instruments ...
First combine template RNA, primer, dNTP mix, and nuclease-free water in a PCR tube. Then, add an RNase inhibitor and reverse transcriptase to the PCR tube. Next, place the PCR tube into a thermal cycler for one cycle wherein annealing, extending, and inactivating of reverse transcriptase occurs.
Microarray RNA-Seq Contains manual curations of public transcriptome datasets, focusing on medical and plant biology data. Individual experiments are normalised across the full database to allow comparison of gene expression across diverse experiments. Full functionality requires licence purchase, with free access to a limited functionality.
CAGE tags tend to start with an extra guanine (G) that is not encoded in the genome, which is attributed to the template-free 5′-extension during the first-strand cDNA synthesis [2] or reverse-transcription of the cap itself. [3] When not corrected, this can induce erroneous mapping of CAGE tags, for instance to nontranscribed pseudogenes. [2]
The three main steps of sequencing transcriptomes of any biological samples include RNA purification, the synthesis of an RNA or cDNA library and sequencing the library. [16] The RNA purification process is different for short and long RNAs. [16] This step is usually followed by an assessment of RNA quality, with the purpose of avoiding ...
SHAPE has been used to analyze diverse RNA structures, including that of an entire HIV-1 genome. [25] The best approach is to use a combination of chemical probing reagents and experimental data. [26] In SHAPE-Seq SHAPE is extended by bar-code based multiplexing combined with RNA-Seq and can be performed in a high-throughput fashion. [27]