The study of the molecular processes of the nucleus and cell function.

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146 views

What mammalian cell lines are easiest to grow and transfect?

I'm working with HEK293 cells, which are very easy to transfect. Does anyone have other suggestions for cell lines with very high transfection efficiency? Ideally the chromosomes would also be diploid ...
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3answers
844 views

Knockdown of long noncoding RNA (lncRNA) - how is it done?

I don't work at the wet lab and don't know all the details about the knockdown techniques. My question is: How lncRNA knockdown is done? For example - you have lncRNA that is functional in the ...
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1answer
645 views

Why is Sanger sequencing inferior for detecting SNPs in cancer cells?

I am familiar with Sanger sequencing, but at the level of an undergraduate. A lecturer of mine tried to describe Sanger sequencing as losing the sequence information in noise when used to detect ...
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1answer
312 views

What is solid-phase bridge amplification?

During Illumina sequencing there is a step called bridge amplification by which DNA is amplified by isothermal enzymes. What is this stage, and how does it work?
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1answer
74 views

How can I clean phenol extracted RNA?

I extracted RNA from skin and quantified it in an Nanodrop machine. The 260/230 ratio was very low and two peaks appeared in the absorbance curve, but the RNA is ressuspended in RNAse free water. How ...
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1answer
33 views

Amount of reverse transcriptase in µg or mM for qRT-PCR

I am trying to calculate a titration amount for a molecule which I would like to use in my PCR samples. Different molecules have different densities so I would like to calculate the appropriate ...
3
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1answer
106 views

High protein turnover rate and protease inhibitors?

I work with mice, and I want to see what happens to some specific proteins in the mouse brain after IL-1b injection (intracerebroventricular). I have a problem: when I measure the mRNA and protein ...
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2answers
4k views

Why is PEG important for efficient yeast transformation?

One way to do an yeast transformation is by using lithium acetate, a single-stranded carrier DNA, and PEG (1). I was wondering why is the polyethylene glycol important for the efficient ...
3
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1answer
131 views

What molecular processes are involved in pseudopodial extension?

I am curious as to the processes and mechanisms involved in the extension of pseudopodia in amoeba. How does the cell know and control the direction and extent of pseudopodia formation at a molecular ...
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1answer
387 views

Manipulation of gene expression using VP16 fusion and engrailed fusion to a transcription factor?

Today, a presenter briefly mentioned that gene expression in sea urchins during development might be manipulated using VP16 and engrailed fusions. On a slide, it said that expression might be ...
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1answer
1k views

How does formaldehyde cause protein-DNA crosslinking?

How does formaldehyde cause protein-DNA crosslinking? I would guess it's because the strongly polar water molecule interacts strongly with polar residues on a protein-DNA complex, and adding a less ...
3
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1answer
95 views

205 nm UV-Vis readings

Typically we determine the concentration of proteins using a 280 nm reading. However, it is reasonable to use 205 nm. I was curious about the effectiveness of this method.
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1answer
225 views

How Antioxidants affects Human Metabolism?

I'd like to know how antioxidants affects the human metabolism and which ones are essential for metabolism processes.
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1answer
216 views

Why does the hydrolysis of ATP increasing entropy increase the Phosphoryl-­‐transfer Potential?

I am currently taking a biology class. I do not understand this concept. I understand that the electrostatic repulsion of the negative charges, resonance stabilization and hydration stabilization all ...
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0answers
45 views

Experience with Detox Cloning

I just saw this paper. A Highly Efficient Molecular Cloning Platform that Utilises a Small Bacterial Toxin Gene I was curious if anyone had experience with this cloning strategy using the IbsC ...
2
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3answers
917 views

Improving Gel Extraction yields

How can I improve my Gel Extraction yields. We use the standard protocol from Qiagen, gel extraction, dissolve in QG buffer at 42C and purify via anion exchange columns. However, with 500 ng we ...
4
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1answer
171 views

What is a simple protocol for staining cells in suspension?

I am an engineering student studying how electric fields affect cells, specifically the phenomena of electroporation in living cells. I know that electroporation is widely used for introducing genes ...
3
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0answers
77 views

How does LCR compare to Assembly PCR

The question pretty much explains itself. How do the two methods compare? I've always used Assembly PCR but the method is prone to mistakes and I'm curious how it compares to Ligase Chain Reaction ...
9
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1answer
114 views

E. coli values for [enzymes], [metabolites] and kinetic rates

In my attempt to create a metabolic model of E. coli, I have found a comprehensive list of metabolic reactions and their stoichiometry. The one I am currently using is E. coli model iJO1366 (more ...
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1answer
82 views

What in soy are people allergic to and will these allergens be passed along if an animal that has eaten soy is consumed?

I have two related questions about soy allergies and possible reactions. What is it that are humans who have soy allergies actually allergic to? What is the allergen? If an animal eats soy, are ...
3
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1answer
77 views

Can I use PCR buffer instead of cDNA synthesis buffer?

I am using Fermentase First strand cDNA synthesis kit but its buffer is over. I need to work today but I have not access to any sorce currently. I have PCR buffer in the lab. Can I use PCR buffer ...
4
votes
1answer
168 views

Can forensic DNA analysis be used to generate a visual approximation of a suspect?

In light of the current US supreme court case, I'm curious if enough information can be teased out of a DNA sample to get a "reasonable" approximation of the suspect (never mind the legality). I ...
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1answer
360 views

How does Topoisomerase II inhibition affect cancer cells?

Topoisomerase II poisons represent some of the most important and widely prescribed anticancer drugs currently in clinical use. These drugs encompass a diverse group of natural and synthetic ...
3
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1answer
451 views

Difference between “transcriptional activity” and “RNA expression”

I am working with a network object where the nodes are different events in molecular biology. Specifically; Catalytic activity of X, RNA expression of X, GTP-bound activity of X, Kinase activity of ...
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2answers
5k views

What is the difference between SOLiD, 454, and Illumina next-gen sequencing?

I've started teaching myself about next-generation sequencing in preparation for a new job, and I'm wondering what the main differences are between the 454, SOLiD, and Illumina/Solexa machines, in ...
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2answers
195 views

Question about equilibrium potential formula

My biology book says, that the equilibrium potential for an ion with a charge of +1 is: $$E_{ion}= 62mV \biggl(\log\frac{[ion]_{outside}}{[ion]_{inside}}\biggr)$$ Where does the 62 mV come from? How ...
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1answer
238 views

Why am I getting low transformation efficiency with DB3.1 E.coli cells?

I am making competent cells using DB3.1 E. coli cells. Even after following the exact protocol (Inoue method for ultracompetent cells) given in 'Sambrook and Russel', I am not getting transformation ...
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2answers
751 views

How do I prepare and clone from E. coli DNA?

I'm looking for a protocol to get genomic DNA from an E. coli sample so that I can clone a small portion of it using PCR into a plasmid. (< 500 bp in this case). It seems OWW (Open Wet Ware) ...
6
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1answer
202 views

Why does not the host produce any immune response to antiserum antibodies?

When an antiserum is injected to a person to protect oneself from a certain disease, the antibodies in the antiserum come from another organism. The question is: Why don't the injected antibodies ...
3
votes
1answer
99 views

What is the extent of the effect of Tris on E. coli?

I was a fool and dissolved my antibiotic (Kanamycin) into Tris Buffer rather than H2O. The Kanamycin still seems to be active but a fellow labmate mentioned that Tris messes around with the membrane ...
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0answers
49 views

Solid phase use in HIV rapid tests

I have another question in regards to my HIV test research. The rapid tests like Orasures Oraquick contains a strip of synthetic peptides that are used to represent proteins found in the envelope ...
2
votes
1answer
165 views

Can Taq polymerase be used instead of polymerase Vent exo (-)?

Instead of using polymerase Vent exo (-), can I go with the usual Taq polymerase? Do the PCR conditions change (the temperature and master mix concentrations ) in the these two conditions? Do the ...
7
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2answers
789 views

Bacterial cell lysis - what solution to use?

I am trying to determine how quickly detergents act on bacterial cells (cell lysis). I would like to compare some detergents at difference concentrations for bacteriolytic activity. I don’t care about ...
10
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1answer
103 views

When running a gel, what could cause a standard to run “faster” than usual?

We are running a gel on the products of a restriction digest to determine the size of an insert. We know the vector backbone size should be ~2.9kb; however, the standard appears to be running too ...
6
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1answer
102 views

Bicoid regulation of hunchback

I'm learning about development via the example of Drosophila embryogenesis. I understand that bicoid regulates hunchback, among other genes. My question whether the regulation is direct or indirect? ...
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1answer
910 views

High Glucose vs Low Glucose DMEM for Cell Culture

I've noticed that in mammalian cell culture, there are often two types of DMEM available. High Glucose and Low Glucose. Does it matter which type I use for culturing of cells (e.g. Hela or HEK293)? ...
6
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2answers
107 views

DAM enzyme distances move along the genome

I am fusing a protein with a Dam enzyme (http://en.wikipedia.org/wiki/Dam_(methylase)). The idea is that when the protein binds to the DNA, the Dam enzyme will start methylating nearby GATC sites, ...
8
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1answer
124 views

How realistic is to use DNA for long term storage?

This is mainly a followup question to the recent paper Next-Generation Digital Information Storage in DNA. Personally, while I agree about the data density of the format, I can't help point out the ...
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1answer
163 views

Is there a program that simulates biology on a molecular level?

Is there a computer program that simulates biology on a molecular level? Software that has rules that simulates the rules of molecular-biology?
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2answers
3k views

DNA as an acid? [duplicate]

Possible Duplicate: Does DNA react in all of the ways most other acids do? Even if DNA is made up of nucleotide bases, it is said to be an acid. Why is this?
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2answers
180 views

Do Yeast Insertion constructs revert?

If I insert a new gene with a yeast integrating plasmid and select with a drop out culture once, can I assume that the newly integrated gene will stay in the strain without putting selective pressure ...
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5answers
273 views

What is the lowest common denominator of cancer?

What is the lowest level attribute that all cancers share? Also, what is the highest level attributes that all cancers share?
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4answers
481 views

How are the boundaries of a gene determined?

What statistical processes and methods are used by geneticists/molecular biologists to know where one gene starts and one ends?
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1answer
78 views

Why would we overexpress Sir2 by overexpressing its hypomorph (dSir2-EP2300) in C. elegans?

Can't we just overexpress regular Sir2 in the paper? Rather than overexpress a reduced-function gene? The paper is Burnett C, Valentini S, Cabreiro F, Goss M, Somogyvári M, Piper MD, Hoddinott M, ...
8
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1answer
98 views

Why is propanoic acid often added to the diet of C. elegans?

And what are its effects on C. elegans? The paper is Burnett C, Valentini S, Cabreiro F, Goss M, Somogyvári M, Piper MD, Hoddinott M, Sutphin GL, Leko V, McElwee JJ, et al.. 2011. Absence of effects ...
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3answers
80 views

Is DNA mutation locally energetically stabilizing the DNA molecule

I am no biologist, but as a physicist, a spontaneous mutation (seen as a chemical transformation) should lower the energy of the system, at least locally. So I wonder if any research has been done ...
8
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2answers
280 views

Do antibiotics attenuate immune response on subsequent exposure to same bacteria?

A healthy immune response to a bacterial infection includes "memory" to permit the body to thwart subsequent exposure to same bacteria. What are the dynamics of using antibiotics on initial exposure ...
4
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1answer
51 views

How to find ion/water channel related genes

We now have a collection of transcripts at hand. We would like to investigate some particular ones, which are ion/water channel related. How to perform this? Could anybody point out how to find the ...
2
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1answer
111 views

In cancer, why do cells duplicate themselves?

In regards to cancer why do cells replicate themselves? If it's a mutation, what kind of mutation would this be classified as?
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1answer
23k views

Absorption ratios 260/280 and 260/230 for RNA

I extracted RNA from different cell lines, an I want to perform reverse transcription and then PCR. To get good results, in which range should the absorption ratios 260/280 nm and 260/230 nm be? And ...