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16

It has been well established that mRNA abundance serves as a poor proxy for protein abundance in most cases. This paper on yeast and this paper on cancer both establish this, although using older techniques (SAGE and microarrays, respectively), while this more recent review discusses the topic in light of more recent technologies (e.g. RNA-seq). Perhaps the ...


15

I don't have any literature to back this up but I doubt that it occurs (at least frequently). For example, imagine a simple three exon gene. Upon splicing exon 1 to exon 3, exon 2 would be excised as part of the intron lariat and subsequently degraded. So in order for exon 2 to be spliced to exon three you would need to either have splicing between exon ...


14

After performing a quick literature search, I am, as with GWW, unable to provide any literature against this occurring, although this paper by Black (2005) states that exons in multi-exon pre-mRNAs are always maintained in order. Black DL. 2005. A simple answer for a splicing conundrum. Proceedings of the National Academy of Sciences of the United States ...


9

See this paper. They have studied RBP-protected sites in the entire human transcriptome by RNA-protein crosslinking followed by RNAse digestion and sequencing: PIPseq. Figure 1 of the paper shows distribution of protein protected sites in RNAs. They also correlate it with different regions of mRNA and its expression. They show number of protein protected ...


7

If the anti-sense transcript is correctly annotated and in the databases (a very very big if), then it will have a different name. For example, the mouse Msx1 anti-sense transcript has the RefSeq accession NR_027920 while the sense transcript of the same gene is NM_010835. In general, each transcript that is transcribed from a given locus has a different ...


7

No, this will not happen. mRNAs are inspected in the nucleus before they are exported into the cytoplasm (at least in eukaryotes), where transcription and translation don't happen at the same place. This ensures that no mRNAs without stop codons or premature stop codons are exported. This phenomenon is called "mRNA surveillance". mRNAs that do not pass this ...


6

Yes, you can find mutations in the genomic DNA which affect splice acceptor sites. Wikipedia lists the following outcome: Mutation of a splice site resulting in loss of function of that site. Results in exposure of a premature stop codon, loss of an exon, or inclusion of an intron. Mutation of a splice site reducing specificity. May result in variation in ...


6

A good question (if a little mixed up on transcription vs. translation!) AUG is not always the start codon, but whatever the codon is it will always code for Methionine (or fMet, but still a variation on Met), even if the codon codes for a different amino acid otherwise. A separate transfer RNA (tRNAi, the initiator tRNA) is used for the arrangement of ...


6

There are some tools for predicting the binding: TargetScan (based on seed match [primary], extra pairing, sequence context 1 — nucleotide composition around the site etc [secondary]) miRanda (based on hybridization stability and seed match[primary] and sequence context [secondary]) PicTar (adds a layer of evolutionary conservation criteria) 1 Context ...


5

If your goal is to avoid regulatory elements, I would not believe any prediction program, since new regulatory elements are found every day. The best way would be empirical and just clone and infect with a mutated 3'UTR and see if your gene's regulation is perturbed. You could at least swap the entire 3'UTR with that from another gene to see if there are ...


5

This study in E.coli is useful in setting out some additional issues at play here, in a system where post-translational modifications are basically a non-issue. The authors see clearly a correlation between mRNA and Protein levels (Fig. 3C) and theorize that differences translation rate and protein turnover are responsible for differences in protein level ...


5

Ribosome doesn't read DNA. Transcription adds several layers of regulation tactics: you can control transcription itself, you can control mRNA. mRNA amplifies genetic information that you need at the moment. Having many copies of a matrix really helps. Archaea have splicing. It's not a good idea to splice your DNA. EDIT per your request of some ...


5

First you have to define what you really mean by miRNA-X targets mRNA-Y. If you mean direct targeting then there are assays to verify it. To verify if the miRNA can potentially target the mRNA independently, what is routinely done is a reporter downregulation assay (usually luciferase). You clone the 3'UTR of your target mRNA downstream of the reporter gene ...


4

The only information you are missing is a way to identify the splice sites. There are many ways of doing what you need. The simplest, assuming you are sure of the origins of the mRNA, is to use a BLAST flavor, either plain BLASTn or, even better, BLAT, to compare your mRNA sequence to the genome of interest. BLAT really should be all you need if the mRNA ...


4

I found the link to a commercial product by Evrogen here: http://www.evrogen.com/technologies/normalization.shtml They claim their method is compatible with nextgen sequencing platforms: cDNA normalization using duplex-specific nuclease (DSN) is a highly efficient approach that can be applied for normalization of full-length-enriched cDNA (Zhulidov ...


4

According to my book Molecular Biology Principles of Genome Function by Craig et al, eukaryotic RNA degradation does not have an initiation, as in bacteria where pyrophoasphate hydrolase hydrolyses the 5'-triphosphate to 5'-monophosphate, and is therefore the initiatior. In eukaryotes the polyA-tail is shortened by a deadenylase enzyme complex which ...


4

mRNA specific queries Add this to your NCBI query: AND mrna[filter] If it's not available in major databases it may not be known. However mRNA data can be obscure to find even in those major databases but it should be there. I would be surprised if that is the case, however since you're not working with model organisms there may well be nothing available. ...


4

Concise Answer The 5′-UTR region of a eukaryotic mRNA is derived from the RNA transcript of the region of a gene between the transcription start site and the DNA corresponding to the translational initiation codon. It differs from that region of the initial transcript in most cases by having a modified guanosine nucleotide added at the 5′-end in a ‘cap’ ...


3

The protocol you are using will not only leave the sample with rRNA but also non coding RNA. Many RNA protocols will separate mRNA by affinity of a carrier to the polyA tail. This protocol references an older paper that estimates that only 5% of RNA is mRNA. I'd be surprised if this ratio changed by more than 2-3 fold in drosophila. I assume that %age ...


3

Shigeta's got a point: the ribosome is latched onto the mRNA so those two are intrinsically linked. You're really asking whether the ribosome comes off first or whether the tRNA does, but it's actually the new polypeptide, which makes sense: The stop codon is recognized by a protein, the polypeptide chain release factor (RF), which triggers the ...


3

There are references in the literature to the phenomenon of "exon scrambling" which seems to be what you are asking about, but the prevailing view is that the evidence for this process, which comes from comparing EST sequences with genome sequences, can be explained by cloning artefacts occurring during EST characterisation. Certainly I agree that there is ...


3

First of all, it is the transcript and not the gene that starts with an AUG. Also, there are actually quite a few transcripts that start with different initiation codons, they are just the exceptions rather than the norm. As for the need, you can think of START and STOP codons as punctuation. The AUG is read like the first (capital) letter of a sentence (...


3

RNA Pol doesn't worry about stop codons. Transcription termination can occur through the formation of a hairpin in the new RNA sequence, or through the action of Rho proteins. A lot of the time prokaryotes have polycistronic mRNAs, that is, mRNAs with multiple protein coding regions. The stop codons are detected during translation, so you will often have ...


3

Point to know : aminoacyl-tRNA binds to mRNA its not just t-RNA.. So if there is no Amino-acid there is no aminoacyl-tRNA of that aminoacid.. so if there is no aminoacyl-tRNA, the anticodon of tRNA doesn't form a bond with mRNA, so the protein production halts (until the Amino acid produced). If the protein is not formed within a certain long time, the ...


3

As far as I understand it (and I'll preface this by saying that initiation is not my strongest point), but prokaryotes utilize the beautiful AGGAGG Shine-Dalgarno sequence. Usually around 8bp upstream of the start codon, it is this sequence that the prokaryotic ribosome seeks out to initiate translation. It does this through a complementary region in the 3'...


3

As pointed out by others this is not true. I just verified this for humans (annotations from gencode21). Methodology: Obtained the start points of the CDS for all genes For each exon of each gene, calculated the distance between the CDS start and exon end Obtained the remainder after dividing this value by 3 (modulo) Commands: Command-1: awk '...


3

Short answer The 5' UTR on the mRNA includes sequence from the Transcriptional Start Site (TSS) to the first exon. Promoters are usually associated with a corresponding TSS. Longer answer In defining a UTR we must consider where transcription begins. Strictly speaking transcription begins at the Transcriptional Start Site (TSS). A useful website for exact ...


2

You should ask this question over at biostars.org -- and could use a bit more clarity. Do you actually have RNA-seq data that you want to use to find the exon/splicing structure of an mRNA? First step would be to use a splicing-aware aligner. A few free ones are: STAR TopHat GSNAP Let us know why that wouldn't do what you want when you re-ask this ...


2

As others have stated, splicing does happen cotranscriptionally but you cannot be actively transcribing the RNA while it is being transcribed. However, it is generally accepted that a transcript can be polyadenylated while the RNA polymerase is still transcribing - this is called the torpedo model. The distinction is that the transcript has been cleaved ...


2

It's slightly more complicated than the comments above indicate. Glover-Cutter, K et al. (2007) RNA polymerase II pauses and associates with pre-mRNA processing factors at both ends of genes. Nature Structural & Molecular Biology 15: 71-78 The authors present evidence that RNA polII pauses 0.5 - 1.5 kb downstream of the poly(A) site where ...



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