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Questions tagged [polymerase]

Enzyme that catalyzes the serial addition of monomers to synthesize a polymeric biomolecule. This term usually applies to DNA/RNA polymerases which synthesize these nucleic acid polymers from mononucleotides.

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Why do the most structurally well-characterized bacterial RNA polymerases belong to thermophiles?

From Lewin's Genes (11th edition, page 515): The only bacterial RNA polymerases for which high-resolution crystal structures have been solved, however, are from two thermophilic bacterial species, ...
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How is the right amount of nucleotides synthesised during homologous recombination?

I'm reading about homologous recombination in the context of double-strand break repair. It seems like the trick is that around the break, homologous recombination uses a template duplex to generate ...
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What makes/breaks the hydrogen bonds between DNA and RNA during transcription?

So I know that RNA polymerase catalyzes the phosphodiester bonds that hold the sugar backbones of a growing mRNA molecule together during transcription. However, I'm less sure about the hydrogen bonds ...
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Can Phi29 polymerase work at higer temperature?

Phi29 polymerase is often used at 25-30 degrees Celsius and heat inactivated at 60-70 degrees. I couldn't find any data on its functionality on temperatures around 40-50 degrees. Before testing it ...
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How does cell detect if a RNA polymerase II is stalled during transcription and in turn deploy the proper transcription-coupled repair factors?

When a segment of the template strand of DNA is damaged due to factors such as UV radiation, a lesion is created that would effectively block the passage of RNA polymerase II during transcription. ...
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How do cells determine RNA types?

I was reading about the different types of RNA polymerases, and I am confused as to how a cell determines which type of RNA it is transcribing. According to this nature article: RNA pol I ...
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Rapid polymerization inside cells

I am interested in (rapid) polymerization or (rapid) long mollecule synthesishappening in the intracellular space. Clearly protein synthesis or DNA/RNA synthesis are examples of elongation of ...
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Terminology of the sequences of promoters in relation to DNA strands

I'm studying molecular biology and I'm trying to understand an experiment which shows the importance of promoters in the relative transcription level (RT). The image below comes from Rolf Knippers' ...
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Do all RNA polymerase in Eukaryotes share the same transcription factors?

I know that TFII proteins bind to cis elements in DNA and help initiate transcription for RNA polymerase II. But do RNA polymerase I and III also use these proteins when transcribing genes? Also, do ...
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Are bacterial RNA polymerases distributed evenly throughout the cell or localized?

Bacterial cells aren't internally compartmentalized with membranes (like eukaryotes). This naturally leads to an image of a homogeneous interior, but bacterial cytoplasm isn't homogeneous. Case in ...
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Why do we use klenow fragment in pyrosequencing?

Why do we use klenow fragment in pyrosequencing instead of polymerase
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Definition of nucleotide-associated proteins and RNA-polymerase associated proteins

Nucleotide-associated proteins and RNA-polymerase associated proteins, I found those two terms in an article but I didn't ...
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DNA replication: How many DNA polymerase molecules work in parallel?

Background research I am aware that DNA polymerase works in pairs, at least. This is to process both opposite stands of a given chromosome. The 3'-to-5' "leading strand", and 5'-to-3' "lagging strand"...
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Estimating RNA concentration from Specific Activity of Labeled RNA

The question reads, Using T7 RNA polymerase to transcribe in vitro a 100-nt RNA off a DNA template. This RNA contains 19 Adenosines. In your 100 uL transcription reaction you added 1.00 µL of α32P•[...
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Why does Taq polymerase add 3' adenine overhangs?

Is there a mechanism for the preference of Taq polymerase to add a non-templated 3' adenine (overhang) instead of other bases?
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Does the assembly of the PLC transcription complex utilize phosphorylation in binding of the GTFs?

in the recruitment of general transcription factors during transcription initiation is phosphorylation utilized?
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Why do you need primers in PCR? [duplicate]

I have read that DNA polymerase requires a primer to bind to the DNA, but I am confused as to why this is the case. When DNA undergoes replication in the cell, there are no primers in the nucleus. Why ...
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What are the reasons which make intron detection uncertain?

Intron/exon sequence detection seems to involve statistical prediction which can at best deliver a guess (until experimentally confirmed) as to where the splice site is. What are the reasons why ...
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Why and how is DNA synthesis so much faster then RNA synthesis in bacteria?

DNA synthesis in E. coli is 20x faster than RNA synthesis at 1000nt/s vs 50nt/s. (Mirkin'05) I find that perplexing since DNA polymerization has better proofreading than the RNA variety, which ...
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Will bacterial promoter without either -10 or -35 sequence allow no transcription?

I have a strong constitutive promoter on a plasmid inside of E.Coli, which expresses a gene. Having either -10 or -35 sequence modified to random sequence, will it ensure zero transcription from that ...
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What happens when we re-start a PCR reaction?

Recently when my PCR reaction was running there was power fluctuation and the entire lab was blacked out for a few minutes and unfortunately PCR that I was running got switched off. So, would it be ...
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Storing Taqman plates with polymerase in fridge before running

I'm going to be running some Taqman assays on cDNA generated from RNA collected from various samples, and will end up running a bunch of plates (right now the setup is 1/2 a plate per sample, ...
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Stop codons and exons?

If we had a hypotheical gene called gene exampleGene and this gene had 5 exons, labeled A, B, C, D, and E in that order on the chromosome, could it be the case that the stop codon for this gene be on ...
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Does DNA polymerase always go the same direction?

I read that mutations are more likely to occur on "the strand that DNA polymerase replicates discontinuously". Does DNA polymerase always replicate the same strand discontinuously and, if so, how/why?
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DNA polymerase in PCR (polymerase chain reaction)

Can the DNA polymerase in PCR (polymerase chain reaction) recognize both DNA and RNA for use as a template? I want to know is it possible if my primers bind to contaminant RNA and then any DNA ...
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Why do some bacteria have most genes on the leading strand of the genome?

Genes in the (+) strand are black and genes in the (-)strand are red. The gene distribution in E. coli genome is somewhat expected: transcribed regions would tend to alternate with non transcribed ...
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How can E. coli proliferate so rapidly?

The E. coli has a genome with approximately 5×106 bp. The main DNA polymerase involved in its chromosome duplication (DNA pol III , the one with highest processivity) can polymerize ~103 nucleotides ...
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Are there U-rich transcriptional pause sites?

I've heard a statement in a talk that U-rich sequences cause transcriptional pausing. The intuitive explanation for this was that the UTP concentration is lower in the cell than the ATP concentration. ...
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A question about IPTG induction

I have found the information: `In the absence of lactose, the lac repressor binds to the operator sequence on DNA and bends the DNA by 40 degrees. This blocks access of T7 RNA polymerase to the ...
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First RNA polymerase-mRNA

We know that RNA polymerase produces mRNA by reading DNA strand. Which enzyme produces the first RNA polymerase if other RNA polymerases are synthesized in the same way like other enzymes(proteins) ?
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How is the transcription direction of RNA polymerase determined?

When transcription factors attach to the DNA strand - How do they know in which direction they have to initialize the transcription by rna polymerase? Is it always read in the same direction anyway? ...
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Why do we need deep sequencing?

Why do we need deep sequencing? Why cannot the sequencing technologies read all the nucleotides correctly at the first read? Sorry since this question is too trivial, I don't have a biological ...
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DNA Replication

I just wanted to understand the basic steps behind the replication of the lagging strand of DNA: Have helicase unwind it first DNA Primase lays down RNA primers in fragments, called Okazaki fragments ...
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Where does Taq polymerase migrate during electrophoresis?

Assuming a 1% Agarose gel with TAE. Follow up questions: Is there a way to stain for it? Could the polymerase be captured and reused?
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Has anyone ever sucessfully translated xRNA or yRNA? [closed]

I've recently been researching the subject of size-expanded nucleobases in alternative genetic sets. Many papers describe the, at least, partial success in replicating xDNA and yDNA, as well as ...
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How are DNA polymerase error rates measured?

It is well known that the first DNA polymerase, Taq, is quite error prone. Newer generation commercial enzymes that have either been isolated from different thermophile species or have been improved ...
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Open protocol for Ligase Independent Cloning

Ligase Independent Cloning is a protocol that allow an insert to be integrated into a vector without ligation. It uses T4 DNA polymerase with only ATP to first chew back from blunt ends to create ...
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number of RNA pol II molecules in a given human nucleus?

Does anybody have any estimate on the number of RNA pol II molecules in the nucleus of a given human cell? For example, how many RNA polymerase type II protein complexes would there be on average at a ...
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Why is DNA replication performed in the 5' to 3' direction?

DNA replication goes in the 5' to 3' direction because DNA polymerase acts on the 3'-OH of the existing strand for adding free nucleotides. Is there any biochemical reason why all organisms evolved to ...