Questions tagged [ecoli]
Escherichia coli, a gram-negative, rod-shaped bacterium commonly found in the lower intestines of warm-blooded animals. Used extensively in laboratory research.
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I accidentally plated DH5a E. coli and left them in the 4℃ for 12 hours, if I put them into the 37℃ incubator will they start growing?
I started ligation of my constructs late last night and was super tired, since I used all of my ligation mixture I would rather not have to re-ligate.
I did my transformation procedure as follows:
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How was Campbell's Model of phage lambda integration into E. coli definitively proven?
In a series of seminal papers summarized in Annual Review in Genetics (2007) 41 1–11, Allan Campbell proposed a model that assumed phage lambda integration in its host (E. coli) genome. This is in ...
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Problems reading my gel electrophoresis result
This is an image of my gel electrophoresis result.
I used the alkaline lysis method to extract E. coli plasmid and put it on gel electrophoresis. Based on what I had learned, I expected to see a band ...
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Correlation of Meselson and Stahl with “multifork” replication in E.coli
Because of the limiting value of the rate of DNA replication, rapidly dividing E.coli use multiple replication forks [1][2]. Thus, DNA replication of one generation has already begun in the previous ...
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What are possible tweaks when a protein expression from a plasmid shipped on paper fails?
In my group we recently tried to express a protein necessary to perform an analytical method in BL21(DE3). The first two attempts did fail and no colonies did grow on the agarose. I hear, that the ...
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Methods of denaturing BL21 E.coli other than using lysis buffer and sonication
The goal is to try different denaturing methods for BL21 E.coli. However, finding detailed paper on this is challenging as most articles do not seem to include the details of their denaturing ...
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Test for e. coli in murky water
We had some tests done on some stream water. In the results report, the lab said:
The coliform/e. coli test was requested and [we] attempted to test for the bacteria but the sample was too dark in ...
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Mutations in a Petri dish overnight
How long does it take for a bacterial culture in a Petri dish to experience all possible single base pair mutations? Can 12 hours be enough?
I want to get an intuition for whether a given mutation is ...
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Why do E.coli form parallel clusters?
Discalimer: I am not a biologist. I am modelling E.Coli growth and in the Wikipedia gif one can see observe some groups where the bacteria are one next to each other, oriented at approximately the ...
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Why do coliform bacteria smell so bad?
I have done multiple cultures of E. coli bacteria and related species over the course of my education. Almost every time, they had this revolting smell. However, other organisms used for genetics ...
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Absent gene expression from one of two vectors co-transformed into the BL21(DE3) strain of E. coli
I've been trying to express two proteins in the BL21(DE3) strain of E. coli.
One gene is in a pCR2.1 vector and the other in a pET-expression vector. When I induce with IPTG and run on an acrylamide ...
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How to estimate mRNA counts from Relative Promoter Units (RPU) or RNAP per second (PoPS) in E. coli?
I need a rough estimate on how to translate RPU or Polymerases Per Second (PoPS) to mRNA count (or even better protein count) in E. coli.
I understand that any number we come up with will be a very ...
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How can I improve efficiency of Ecoli transformation?
I am an intern in biology institute. I have a 17.3kbp plasmid need to transform to Ecoli. But I have tried many time but have no or very few colony on LB plate.
I know large plasmid have less ...
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Why pellet and resuspend E. coli for plasmid prep
For maxipreps, cant you just add all the stuff that would be in P1 (RNase A, EDTA) then just add P2? Because if we pellet cells then resuspend theres got to be a reason, right? Is it slats and other ...
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What happens when the bacteria in the petri dish lack nutrients for long time?
If I put some E.coli in a petri dish filled with limited nutrition in room temperature and forget them for months or years. What will happen in the final after they overgrowth and used all nutrition ...
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Low cost cell free expression system
I am a graduate student looking to incorporate some protein engineering into my training. I have been reading a lot about E coli lysate based systems, but they still require 3rd party additives like ...
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Did early 20th century researchers state why they used E. coli as a model organism?
I am investigating the early uses of E. coli as a model organism. Sadly, many early 20th century papers are not in English. In those that are, I have been unable to find explicit statements as to why ...
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E. Coli metabolization of paracetamol
For my science fair project, I did an experiment on how paracetamol and ibuprofen affected the growth of bacteria. By day 5, however, parts of the paracetamol blackened (image below). I am wondering ...
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E.coli fermentation - process flow
I have an engineering background and have been assigned a project to look into to failures during E.coli fermentation process using the data available before/during/after the process
I have learnt ...
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Dissolving cell pellet after sonication
I was doing a protein prep and I made a mistake. After sonicating my cells, I was supposed to centrifuge and collect the supernatant (my protein is soluble and comes in the supernatant). I however, ...
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Are there ways to genetically increase mutation rate of E.coli?
Are there ways to increase mutation rate of E.coli using genetic modifications? I know possible ways it can be done without genetic modifications by exposing the cells to stressful conditions like ...
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Plasmid specific DNA degradation
I'm interested if there is a way to degrade the plasmid DNA inside an E.Coli cell specifically so that the method does no harm to the chromosomal DNA. First I was thinking about restriction ...
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Why do two different E. coli reference genomes have different lengths?
I've downloaded two different reference genome of E. coli (E. coli K-12 MG1655: U00096.1 and E. coli K-12 MG1655: U00096.2) and they have different lengths. I searched for the meaning of reference ...
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What nutrients are best suited for growing E.Coli
I am looking to grow E.Coli (In a nutrient agar dish) to be used in an E.Coli lawn and was wondering what specific nutrients should be used to ensure the E.Coli grows optimally? Any answers or links ...
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How does a bacterium age without cell division?
If somehow a bacterium, let's say E. coli, is engineered in such a way that it loses the formation of ftsZ ring (or other ways that inhibit cell division), and if other conditions are optimal for ...
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Codon tables and the wobble hypothesis
In E. coli, there are only 47 different tRNAs but 61 potential anticodons. This is because, from what I understand, the third base of the anticodon can pair by wobble rules.
However, it is known that ...
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What is the mass of an E.coli cell?
I'm trying to get into Biology, reading introductory texts. A discussion on E.Coli estimates the mass of the cell from $$density =1g/mL=1g/cm^3$$ and $$volume=1µm^3$$ From this I get $$ mass=density × ...
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Unwanted E. coli product in plasmid product
Using a TOPO XL Cloning kit, our lab took some amplified human DNA, inserted it into the vector, and attempted to clone it in E. coli. However, upon sequencing the purified plasmid product, the ...
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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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Relative densitometry from SDS PAGE
I'd like to perform densitometry on a Coomassie stained SDS PAGE gel to compare a recombinant protein's expression levels under two conditions. I'm using BioRad's Image Lab software.
My questions are ...
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Is it possible to grow E. coli K12 with algal amino acids as the sole carbon source?
Instead of glucose or glycerol, is it possible for E coli to grow on an amino acid mixture as the sole carbon source and, if so, what should the concentration be?
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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 is the spacing between stop codon and transcription terminator?
When one tries to express a protein from a synthetic plasmid in E. coli, how many base pairs should there be between stop codon and the transcription terminator site?
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Can I leave BL21(DE3) cells in room temperature?
I am preparing competent cells, and I finished inoculating a single colony in SOB. It has been incubating at 37 degrees Celsius for almost 16 hours since, and it's getting very late where I live. I ...
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Do I need to serially dilute E. coli cultures for optical density measures?
I am expected to track cell growth by measuring the incubating culture's cell concentration every 30 minutes or so. So my questions are: Why do I need to do serial dilution (assuming that I do)? Why ...
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When do plasmids replicate relative to its host cell cycle?
For plasmids is so much shorter than their host cell's genome (about 1/1000 in my case), it will take only 1/1000 time for it to replicate.
With respect to cell cycle, when will that replication ...
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Where to find E.coli gene expression data?
I am searching E.coli whole genome expression data with different conditions, any suggestion is appreciated. Condition could be for example different growth temperature, different medias, etc.
I have ...
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How to prevent e coli from clumping (for FACS)?
I'm performing FACS on e coli, but the cells are clumping together so each event is multiple cells.
I ran a control where I had one flask of e coli expressing GFP, and one flask expressing RFP. Run ...
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Exact location of E. coli in human body
E. coli lives in lower intestine. But exactly where can we find E. coli, and where is there none (or a really small amount) of it?
I need the exact names of parts of the lower intestine where we can ...
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Recombination frequency as a function of genomic location in E coli?
Are there any databases or papers that give recombination frequency as a function of position in the E coli genome?
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Need for two oxygen sensors in E. coli
E. coli has two oxygen sensors: FNR (fumarate-nitrate reductase) and ArcBA (Anoxic Redox Control, two component control systems). FNR directly senses the oxygen, while the interaction of ArcB with ...
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Plotting metabolic networks?
I need to draw a map of the core metabolism of E.coli. Associated with each reaction in the map I have a number that indicates the flux through this reaction. I want the map to reflect these fluxes ...
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What does it mean to perform in vitro experiments with mutant bacteria?
I came across the following sentence while reading the paper:
Nanchen, Annik, et al. "Cyclic AMP-dependent catabolite repression is the dominant control mechanism of metabolic fluxes under glucose ...
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What is the mechanism of oxygen uptake in E. coli?
How does E. coli uptake oxygen? Most of the literature I found is concerned with response to oxygen level supplied in the medium, as opposed to how much is actually transported inside. Can they shut ...
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Consumption of NAD+ in glycolysis
Out of 10 steps in glycolysis, only one reaction- Glyceraldehyde 3-phosphate (G3P) to 1,3-bisphosphoglycerate (PGP), uses NAD+ and thereby producing NADH. Furthermore, this very step is solely ...
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Anaerobic respiration choice in E. coli
Under anaerobic conditions E. coli has two options to generate ATP: fermentation (substrate-level phosphorylation), and respiration (proton gradient, chemisomotic phosphorylation). Which is favored? ...
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Regulation of Cra protein level in E coli
Catabolite Activator/Repressor, Cra protein (formerly known as Fructure Repressor FruR) plays a significant role in central carbon metabolism of E coli. Its activity is inhibited by fructose-1,6-...
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Problem with bacterial transformation with electroporation
I have a problem with a bacterial transformation of a yeast gene that I can not solve.
I isolated yeast DNA and did a PCR to get my product. I am using pCGCUm vector with a GFP construct. I digest ...
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explanation of conjugation experiment in bacteria
In this paper (On the expression of a structural gene) I am confused about what is being plotted in Figure 6. The x-axis contains the fraction of radioactive day and y-axis contains the enzyme ...
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How does blue white screening work in selecting E coli that have taken up a plasmid vector?
The plasmid vector I am referring to is pCR 2.1 - TOPO. I added the vector to the E coli and plated them up onto LB+amp+X-gal plate, then incubated. After incubation the plates had two types of ...
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