histone - H3K36me3 role in RNA polymerase II elongation

I found a paper describing the association of RNA polymerase II with the histone methyltransferase Set2 that methylates H3K36.At the end they say: While our results strongly implicate K36 methylation as having a direct role in RNAPII elongation, the precise role is currently unclear. We propose three possibilities: (i) K36 methylation generates a chromatin structure that is more permissive for RNAPII passage. In this scenario, the loss of Set2/K36 methylation would result in a chromatin structure that is more difficult for RNAPII to pass, whic...Read more

rna - Why does a cell need siRNA?

My question is about small non-coding RNAs. miRNA and siRNA are both known for mRNA regulation. miRNA can both transcriptionally inhibit and degrade mRNA by cutting it. For inhibition just a small fragment (seed) of miRNA is complementary to the mRNA while for degradation miRNA and mRNA must reach 100% match. siRNA is almost the same molecule and its only function is to 100% match mRNA and degrade it. So if siRNA appears to be just a subtype of miRNA, can we confirm that? Or are there any other differences that make these two molecules differen...Read more

Why do we use cRNA instead of initial extracted RNA in the microarray technique?

I am recapping for the exam in transcriptomics and stumbled into a question about microarray. So, our regular workflow is toextract RNA from cellsgenerate cDNA from the initial RNA using reverse transcriptiontranscribe cRNA from cDNA label cRNA with biotinfragment biotin labeled cRNAhybridize to a plate, scan and quantitate So the question is - since we use RNA for the further quntification steps why should we perform reverse transcription+transcription to produce cRNA while we could just use the initial RNA? Is the cRNA more stable than RNA? O...Read more

nucleic acids - Can I ask tip for tissue measuring(especially for RNA preparation)?

I extracted RNA at mouse liver and human blood, but degradation occurs. I think, at mouse liver, too hard to measuring 50mg liver(I am newbie for animal tissue preparation) so spend several minutes. liver is soft but very small, and too hard to keep the weight. (human blood spend time for defreeze, but It degraded, too.)for example, I want to weight 20mg mouse liver, 1. cut liver by scissors about 10mg after set zero point of scales with ep-tube. 2. cut liver almost 10mg, but it's exceed weight 2~3mg, so I have to cut it 2~3mg. 3. this scale is...Read more

rna sequencing - Finding Rna-Seq dataset for C. Elegans (Caenorhabditis elegans)

I want to find a RNA-Seq dataset for C. Elegans to use it in RNA-Skim software. For those who does not know RNA-Skim let me explain it a bit. It is a tool which is used to quantify RNA-Seq data using sig-mers(special kind of k-mer). Firstly RNA-Skim clusters protein coding genes and find substrings that are special to cluster of that gene. After this preperation stage, it takes an RNA-seq data as an input and tries to quantify transcripts in it. So, I need to find a dataset in fasta file format for this step. Since I'm pretty new in this area, ...Read more

Current state of direct RNA sequencing

I had a colleague ask me recently whether mRNAs could be sequenced directly. I found this Nature paper[1] published by Helicos in 2009, in which they describe their developments in the area. It's been several years since this was published, and yet RNAseq and most other transcriptomic analysis is still done by first reverse transcribing RNA into complementary DNA.What is the current state of RNA sequencing? Has Helicos failed to deliver on their technology? Have any other platforms risen to the challenge (or attempted to do so)? Or is the techn...Read more

gel electrophoresis - Alternative protocol for evaluating RNA integrity, using a bleach gel

I hate having to run MOPS PFA agarose gels with DEPC and everything just to verify my RNA integrity. I came across an alternative protocol that uses ordinary household bleach in the gel to inhibit RNases and was wondering if anyone here has tried it and what the success was like? Or any helpful insight is welcomed also.here's a protocol I put together based on what was done in that paper.For a 50ml gelIN a RNase-free 250ml Erlenmeyer flaskMix:5 mls of 10x TAE that I gave you42.5mls ultra purified water0.5g of the agarose I gave you2.5mls of hou...Read more

pcr - RT-qPCR for RNA quantification

I'm really new to RT-qPCR and probably I've just a minor problem than I think.So, what's my problem about.I want to quantify a virus by a specific area within a gene on its genome. I've specific primer which are working fine with the extracted virus rna from my samples. But now I need a template for a dilution series to quantify this virus. And there's my little problem. And maybe you can help me.So I know that the genome of my virus is a negative sense single stranded RNA, so -ssRNA, and moreover I know the virus strain and the gene, which RNA...Read more

transcription - RNA polymerase and DNA helicase

Is DNA helicase or RNA polymerase responsible for breaking the hydrogen bond between the 2 strands during transcription for eukaryotic cells? My textbook (WJEC Biology for AS level) says it is DNA helicase that breaks the hydrogen bonds while RNA polymerase catalyses the formation of bonds between RNA nucleotides. The enzyme DNA helicase breaks the hydrogen bonds between the bases in a specific region of the DNA molecule. This causes the two strands to separate and unwind, exposing nucleotide bases. The enzyme RNA polymerase binds to the temp...Read more

rna - Do ribosomes read mRNA?

So I understand that tRNA bonds to a codon (with an anticodon) in the translation process. I read in my biology textbook that the ribosomes "read" the mRNA strand.Why do the ribosomes need to read the mRNA strand? Are the ribosomes able to intepret the message and bring the correct anticodon to it? Or is the main function of the ribosomes just to join amino acids to make polypeptides? I'm just confused why the ribosomes read the mRNA strand when it seems like there is no need to....Read more

rna - Why is an initiator tRNA required, distinct from the methionine tRNA used in elongation?

I'm confused by why there is a need for different tRNA-methionine complexes for translational initiation and elongation. This paper mentions that It is important that each type of methionyl tRNA be restricted to its separate function, as competition for tRNA by the initiation and elongation machinery could lead to serious problems for the cellI don't understand what these problems may be....Read more