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Questions tagged [rna-sequencing]

Determining the sequence of RNA. This technique is used for discovery and quantification of mRNAs and ncRNAs.

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How can MMLV reverse transcriptase add cytosines to the 3'-end of cDNA

I am working on the mechanism of SMART, a kind of reverse transcription strategy. However, I am confused about the terminal transferase activity of MMLV enzyme. It have been reported since 1998, ...
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How to differentiate transportable element and non-coding RNA in genome?

I am searching for unannotated non-coding RNA in my genome at the same time i want to omit the transposable element from my genome. My sequencing libraries are paired-end and i am searching in the IGV ...
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Why do the RNAseq reads map to only a certain region of a viral genome?

I am looking at RNAseq data and mapping them to 3 RNA segments from Cucumber Mosaic Virus. I trimmed the adapters from the fastq files and converted them to fasta, which I searched against a virus ...
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identification of differentially expressed genes in RNA-seq analysis [closed]

I am using four different packages (viz. EBSeq, DESeq2, edgeR, LPEseq) for identification of differential genes. Now I am confused whose fold change value should I take for further downstream ...
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Is there a data base, tool or method I can use to find out which of my genes code for cytokine receptors?

I have a list of over 600 differentially expressed genes from my single cell RNA seq data analyses. I want to proceed to find out which of my genes code for cytokine receptors so that I can show on a ...
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120 views

Sequence of ribosomal RNA

Is it possible to sequence rRNA directly, that is, using the ribosome rather than the DNA from the nucleus? For example, this paper, Complete nucleotide sequence of a 16s rRNA gene from E. coli, ...
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148 views

Iron metabolism in the brain

I did an expression profiling from publicly available RNAseq data for mouse tissues. While for liver and testes I am getting expected proteins in the top highly expressed mRNAs, for brain I am getting ...
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190 views

What do HG and NA mean in Geuvadis project RNAseq sample labels?

I'm looking at RNASeq Data from Geuvadis website e.g. the file GD660.GeneQuantRPKM.txt.gz. The samples are labeled by e.g. HG00105.1.M_120209_7 or NA20812.2.M_111216_6 What do HG and NA mean? Are ...
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Why analyse transcriptome instead of proteome?

Analysing the transcriptome (RNA-Seq, microarrays, qPCR etc) is probably the most widely used technology to assess gene expression and dynamic cellular processes. The results are then extrapolated (...
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223 views

Which information can be extracted from time course RNA-Seq Data?

So I am very new to the area of biology so I am sorry if this is a stupid question. I have RNA-Seq data carried out over a span of 100 days and I have gene expression data in the following format. ...
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How many samples to run an association study on B-Cell receptors

We are trying to find biomarkers on the B-Cell receptor VDJ regions. To do this we plan to run Single cell RNA-Seq (using the solution from 10x genomics https://www.10xgenomics.com/solutions/vdj/) to ...
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22 views

Metatranscriptomics Extraction

I am attempting to perform metatranscriptomics analysis of a cleanroom. The DNA input is known to be rather low (<1pg). Despite this, I want to still attempt to try it. I am thinking it might be ...
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58 views

If a person is infected with an RNA virus what percentage of the RNA in the blood of the person contains RNA of the virus?

Karius uses DNA sequencing of everything that's contained in a sample to detect all bacteria and DNA-viruses in the sample. Given the way that costs of sequencing are falling this approach seems to be ...
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Why did protocell division coincide exactly with RNA replication?

I know that, in the RNA world, it has been suggested that the first protocell formed by a hollow liposome structure encapsulating an RNA molecule. However, from then on it becomes confusing: I've ...
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Correct structure of RNA [closed]

Of the 2 figures, which is the correct structure of RNA, This is the link : 1: https://i.stack.imgur.com/IYgyv.jpg Sorry, the figure is not to scale. If there's any problem with the question ...
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RNA contains Uracil in place of Thymine [duplicate]

Thymine does not present in RNA but DNA containing thymine why
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19 views

dbSNP clarification

I am currently going through this paper - link. In their methods section the authors have described they have used dbSNP for checking biallelic expression of genes. I would like to know if dbSNP ...
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Should I use glycogen in isopropanol to precipitate RNA or does glycogen have harmful consequences on downstream protocols such as RNA seq?

I have heard that one can use glycogen in isopropanol to better visualise the RNA pellet after centrifugation. What are the advantages and disadvantages of adding glycogen in isopropanol for RNA ...
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176 views

Should I use RNAseZAP on my bench and pipettes before RNA extraction?

Should I spray RNAseZAP on my bench and pipettes before working with RNA? I am regularly using RNase at my bench and would like to work with RNA as well. Could traces of RNase on my equipment degrade ...
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81 views

Where is DNA during the process of RNA extraction using Trisure?

Where is DNA during the process of RNA extraction using Trisure described here? Given that DNA is also a nucleic acid, should I expect the isolated RNA to also contain DNA? However, Bioline claims RNA ...
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184 views

Is DNase treatment necessary before RNA sequencing?

Is it necessary to treat total RNA with DNase before sequencing it? In particular, if the library prep relies on a poly-A enrichment, is it necessary to remove DNA, knowing that genomic DNA does not ...
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67 views

de novo assembly applications

Hey I hope this question supposed to be in this forum. I'm not from the biology community, I'm from the data security community so im not familiar with the words you might use for answers I need to ...
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160 views

the meaning of “Non-primary hits” and “tags”

Can I ask you to help understand the meaning of "Non-primary hits" and "tags" in the following paragraph related to RSeQC: The result table reports total number of reads (excluding nonprimary hits) ...
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RNA extraction from soil

I have been trying to extract RNA from soil without success. I have used commercial kits (The Plant DNeasy kit) because I did not want to buy a complete new kit (Soil) to do 5 extractions. Is there ...
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70 views

The advantages of using a strand specific cDNA in RNA-Seq?

May I ask what are the advantages of using a strand specific cDNA library in an RNA-seq experiment? as far as I know it can make us able to read the code region but what about other advantages?
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56 views

Is my RNA-seq experimental design correct to use it for SNP calling?

I am a newbie here and would highly appreciate your advice about one particular experimental design. We have data from RNAseq experiment which was originally designed to assess differential ...
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2answers
859 views

Biological meaning of read length

I have some FASTQ files in two datasets which are sequences from 16Srna region. The first dataset is amplicons form V4 region and the second is V3-V4 region. However all the reads are 250 nucleotides ...
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156 views

16s rRNA Sequencing From Gut Microbiome (stool)

Do I extract RNA or DNA from gut microbiome (stool samples) if want to do 16s rRNA sequencing?
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1answer
155 views

Do all microRNA isoforms need to be known and sequenced to obtain microRNA expression?

From what I understand, microRNA are short "hairpin" shaped nc structures. When sequencing and counting miRNA, isoform miRNA are "found" which are subsections of varying length and possibly with some ...
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When to decide that two similar sequences are different genes or not?

Working on RNA-Seq data directed me to ask this question. In RNA-Seq jobs, after de novo assembly we have lots of transcripts with different rates of similarity (0 to almost 100 percent) that we ...
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2answers
3k views

How were the first primers made?

I keep reading about how primers are useful in pcr -- they allow you to select a specific dna region. Similarly, in NGS or Sanger sequencing they give you a starting point. The primers I see are about ...
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1answer
229 views

Which are the best programs to analyze circular RNA?

Hello I'm just started with bioinformatics and I would like to know which programs are best for the task of finding circRNA (circular RNA) in sequences of tissues and different cellular types. I have ...
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1answer
254 views

Multiple transcripts matching same gene in de novo assembled RNA-seq data, but FPKM values vary?

I have a data set of de novo assembled RNA-seq datasets across different sample types. When BLASTing, many of the matches of the individual transcripts match to the same gene on the reference genome. ...
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RNA seq: Should non-infected units be used for RNA-seq?

I am designing a RNA-seq experiment to study transcriptomic profile changes upon parasite exposure. Some papers I read, mostly extract RNA from the entire treatment group (organisms exposed to ...
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341 views

What is the meaning of the word 'bin' in the context of RNA-Seq?

I have a question from a book about RNA-Seq. I would like to know the meaning of the word "bin" in the below cited paragraph: RseQC has several nice features not found in the other programs: (a) ...
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1answer
96 views

correction of read counts for spikes

I have 4 RNA-seq samples (1st is control and the other 3 are treatments) and 5 spikes. I used different concentrations for 5 spikes but over different samples, I used similar concentration. for ...
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Blastx: from gi to description

I have done a blastx and use the outfmt 6 option, because I want to work in the tables format. When I do this I get subject ids of the type sp|Q7XJS0|ASHR1_ARATH, sp|Q39547|CUCM1_CUCME, et cetera....
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How much tissue is required to do RNA-seq analysis on a single organism?

How much tissue would be required to do RNA-seq analysis on a single organism? More specifically, if a person wanted an RNA-seq analysis of expression for a single organ, how much tissue would they ...
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1answer
116 views

How do I read an RNA expression pattern?

When reading about diseases one can find links to proteins and their associated genes, an example of which is here. I'm wondering how to decode/read the following graph as a non-specialist in this ...
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1answer
40 views

What determines the amount of data you would get from a sequencing experiment?

When someone performs a sequencing experiment, what determines how much sequencing data you get from the machine (Illumina, PacBio, Nanopore, etc). Or maybe, put differently, how does the machine know ...
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1answer
62 views

where i can download reference MSU7 rice genome?

I am working with RNA Seq data analysis. I have to download MSU7 rice genome. somehow i downloaded from "http://rice.plantbiology.msu.edu/pub/data/Eukaryotic_Projects/o_sativa/annotation_dbs/...
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1answer
55 views

What factors should I consider when selecting a reference genome for mapping?

I am under the impression that the most recent reference genome is typically the best case. What other things should I consider when selecting a reference genome? For example, is there any particular ...
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4answers
352 views

Tools to analyze RNA-seq data

I hope this is a good place to ask such question. I have to do some data analysis on RNA-seq data from human cells. I am currently searching for tools to help me with that. Specifically, I would need ...
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1answer
59 views

How do you obtain specific mRNA transcript levels for comparison from a Hi-Seq dataset?

I have HiSeq data from mice exposed to two conditions. I would like to answer the following question: "Is there a significant difference in mRNA transcript levels when comparing condition A to ...
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2answers
73 views

Is it possible to identify a particular gene sequence from next gen sequencing techniques (especially RNA-seq )?

I have to check the expression of a gene in a fish whose sequence is not known in the fish in question. Sequence is known in an another fish (zebrafish) but the gene has 10 isoforms. The genome of ...
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0answers
51 views

RNA-Seq library normalization and experimental setup

I'm working on an RNA-Seq project and I am trying to figure out library normalization. I'm aware of using the geometric means (e.g. cuffdiff) of the fpkm for the normalization. However, I was ...
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1answer
95 views

Do house keeping genes vary across tissue

I am working on an RNA-Seq project, and I am aware that some researchers use housekeeping genes as a method of normalization. My project has several different tissues, and I was wondering if ...
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43 views

Is it okay to scale the RNA-Seq counts after log-transformation?

On an RNA-Seq dataset, I want to apply a clustering algorithm which requires zero-mean uni-variance gene expression levels. I am wondering whether it makes sense to use "scale" function in R after ...
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120 views

What is the another use of de Bruijn graphs in bioinformatics except DNA assembly? [closed]

I implemented own generic de Bruijn graph, which I use for DNA assembly (alphabet: A, C, G, T). I try to find purpose of de Bruijn graph for another bioinformatics problems, if some exists. I want use/...
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1k views

What is the most appropriate way to normalize gene expression data?

This question comes because reading a paper about normalization of gene-expression data, is not clear if the method for normalize the data is just for RNA-Seq data or could be applied also for ...