Questions tagged [molecular-biology]

The study of the molecular processes underlying life.

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4
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1answer
685 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 ...
9
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1answer
149 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 here)...
1
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1answer
108 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 those ...
3
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1answer
118 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 ...
6
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1answer
1k 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
2k 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 X,...
10
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2answers
23k 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 ...
4
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1answer
833 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 ...
6
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2answers
2k 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
542 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 ...
2
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1answer
376 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
3k 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
303 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
228 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? ...
6
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2answers
150 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
168 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 ...
7
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2answers
54k 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?
4
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2answers
1k 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 ...
23
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4answers
1k 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?
5
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1answer
108 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, ...
7
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1answer
129 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 ...
2
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3answers
126 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 ...
5
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1answer
64 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 ...
11
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1answer
58k 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 ...
8
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1answer
1k views

How fast do different organs turn over cells?

It is said that the human body turns over all cells or molecules in 7 year cycles. This is not quite correct, because there are different organs changing at different speeds. My interest is what is ...
1
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2answers
704 views

Do the simplest bacteria have ribosomes and helicase?

I find it interesting that all life on earth use DNA. I've seen video on how helicase and ribosomes work together to copy DNA sequences (to RNA) with helicase then recreate them using ribosomes. Does ...
4
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2answers
150 views

How do proteins and genes participate in learning?

I am a computer scientist that studies biology and bioinformatics. In the last weeks, I have been trying to study new research directions, and I would like to deepen my knowledge on the role and ...
10
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1answer
6k views

Is the eukaryotic nucleus composed of a single or double membrane?

I know that it is usually considered a double membrane like those surrounding mitochondria and chloroplasts, but I read a review that stated "according to topological details it is actually a single ...
6
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1answer
1k views

How Do Adherent Cells In Culture Attach To A Plastic Dish?

I am particularly interested in MC3T3-E1 cells (mouse fibroblasts), which are adherent cells. Are hemidesmosomes involved in anchoring the cells to the plastic dish? What (if any) other molecules are ...
1
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1answer
86 views

If inhibiting S6 kinase decreases protein translation, then could inhibiting S6 kinase could possibly slow down long-term potentiation in neurons?

From http://en.wikipedia.org/wiki/P70S6_kinase... Phosphorylation of S6 induces protein synthesis at the ribosome. P70S6 kinase is in a signaling pathway that includes mTOR (the mammalian ...
6
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1answer
1k views

Does anyone have any TOPO directional cloning tips? [closed]

I'm just about to start working on a TOPO cloning after I couldn't get it to work with standard restriction/ligation. Does anyone have any tips for TOPO cloning?
7
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1answer
5k views

Purpose of poly(A)+ RNA?

I am learning RNA-seq analysis. I always encounter this phase "poly(A)+ RNA". After searching, I got this: "Most messenger RNAs contain a poly(A) tail, while structural RNAs do not. Poly(A) selection ...
5
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2answers
3k views

Primer design for introduction of restriction sites flanking a gene of interest

I am wondering what the correct method for primer design to introduce restriction sites. Specifically between two methods. 1) Primer first partially hybridises to the gene, has a mis-match where the ...
10
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2answers
1k views

What is the highest competency possible for E coli?

I am looking to find a highly competent E coli strain. I am making a library of a ~6.6kb plasmid and I am not getting high enough efficiency. Does anyone have a suggestion of a strain/protocol with ...
10
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4answers
302 views

How can I avoid digesting protein-bound DNA?

I'm interested in sequencing and analyzing the bound DNA, and minimizing the amount of unbound DNA that gets sequenced through digestion. When digesting protein-bound DNA, is all of the unbound DNA ...
7
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1answer
779 views

Effect of histidine on the binding affinity of HisP

I was asked the following question by my teacher: A gene regulatory protein called HisP regulates the enzymes for histidine biosynthesis in the bacterium E. Coli. HisP is a protein whoes ...
10
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1answer
1k views

How are there alternative initiation codons?

According to wikipedia and the original complete sequence of the K-12 genome, there are multiple non-AUG start codons such as GUG and UUG. How is this possible? I'm particularly curious about the ...
10
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2answers
261 views

What is the least costly method to generate sequential amino acid deletions?

I'm looking to generation sequential deletions from a gene of interest. The total size of this region is 8 amino acids. I'm trying to determine which portion of this region is necessary within the ...
25
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1answer
2k views

ATP cost for gene expression

How would you estimate the number of ATPs required to transcribe, export and translate a single eukariotic protein?
8
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1answer
178 views

PDB Mining: Why Do I Find Atoms Less than 1 Angstrom Apart?

I am attempting to find potential Hydrogen bonds between Hydrogen donors and aromatic ring acceptors. I do this by predicting the location of Hydrogens on residues and then calculating how far these ...
8
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2answers
2k views

What is causing my problem with very low yields when isolating a 42kb yeast plasmid?

I have to isolate a large plasmid from yeast and transform it in E. coli. After transformation, I often get no colonies. One reason for that is the yeast mini prep hasn't worked or the DNA ...
7
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1answer
1k views

What is the mechanism of transgene integration (from expression vector to the host genome)?

How does a transgene (in vector) integrate to the host genome? (e.g. in glass bead method, neither biolistic nor agrobacterium). I already cut some parts (NdeI-PciI) of the vector (pUC18) and ...
18
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1answer
313 views

What is the prehistory of amino acids in cells?

As a followup to Why 20 amino acids instead of 64? and What is the smallest number of amino acids required for life?, I am trying to understand the prehistory of amino acids in cells. All living ...
15
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2answers
721 views

Intrinsically disordered proteins as potential drug targets

Intrinsically disordered proteins (IDPs) are a class of proteins that do not adopt a stable secondary or tertiary structure under physiological conditions in vitro, but still have biological functions....
10
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2answers
5k views

How do I clean and calibrate pipettes, and how often should I do it?

I work in a lab where all the pipettes are shared. We often have visiting students who come and use the pipettes for a short project. So when I work with them, they might have been handled by other ...
6
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1answer
316 views

In C. elegans, why does knock-down of cco-1 in some tissues increase lifespan, and knock-down of cco-1 in other tissues decrease lifespan?

Full question: In C. elegans, why does knock-down of cco-1 via RNA interference in specific tissues like body wall muscle decreases life span, whereas knock-down in the nervous system and intestine ...
8
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1answer
633 views

How do you knockout an E. coli gene without disrupting the rest of the gene cluster?

I'm familiar with the method to build Keio collection using single-gene deletion with an antibiotic cassette. However, what happens when there are gene clusters or overlapping ORFs and you still only ...
2
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1answer
106 views

Is the protein in teardrops still attached to cells, or is it released and free-flowing?

A ScienceDaily article says that the protein in teardrops can kill bacteria. But how does it reach the bacteria?
6
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2answers
1k views

Reverse transcription PCR optimization

What is the ideal amount of RNA to use for the RT? and how much cDNA to use then for the PCR? I did RT with a solution of RNA of 0.36 ug/ul. Then for my PCR I used 1 ul of the cDNA obtained and used ...
6
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4answers
6k views

What are good resources for self-learning modern molecular biology concepts? [closed]

My learning of molecular biology ended in the early 90's (and with early 90's era information). While I don't aspire to be a molecular biologist, I do aspire to better understand modern approaches ...