Questions relating to protocols, procedures, and good practice when using laboratory equipment.

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2
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2answers
122 views

Extending a small fragment of DNA

Is there a way to extend a small fragment of DNA, say 150 bp, by making copies of itself and attaching each copy of that small fragment to the end of that 150 bp sequence? For example, I want a 1 ...
0
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0answers
8 views

Column chromatography protocol for Alternanthera sessilis to fraction phytochemicals [migrated]

I'm in need to find a column chromatography protocol to fraction phytochemical compounds particularly polyphenols in Alternanthera sessilis Red. I can't seem to find one. Plus this is slightly out of ...
0
votes
1answer
19 views

Random Mutagenesis vs Directed Evolution as Strategies to boost expression

Do people use random mutagenesis (say using UV) to generate host variants that have high expression of a metabolite / enzyme? I've seen it mentioned as a strategy but it confuses me as to why. How ...
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vote
1answer
16 views

Voltage sensitive dyes technique: What is the underlying measure?

I just discovered voltage sensitive dyes technique: first of all what imaging techniques do we use? And I have seen that figures are labeled with ΔF/F0, what does it stands for?
6
votes
1answer
89 views

Can microdialysis be made in Drosophila melanogaster?

I've asked this before in stack overflow's cognitive science community, and someone recommended me to ask here: I've found a couple of studies using microdialysis on insects, but didn't found any in ...
1
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1answer
54 views

Does adding antibiotic after 5-10 mins of innoculation affect the protein yield or growth?

I've asked a lab colleague the same question. She said, it would loosen the bacterial cells in the LB medium and plasmids would come out. Is that true? and why?
2
votes
1answer
37 views

Optimizing Gel Electrophoresis: Ampere, Volts and Buffer concentrations

I am a master student in biochemistry, and I have used gel electrophoresis many times before. What I want to know is how one should adjust the mA (mAmpere) compared to the voltage and the buffer one ...
6
votes
2answers
222 views

Measuring fitness / lifetime reproductive success (LRS) in Drosophila

I am planning a fitness assay of Drosophila melanogaster. I'd like to get a good measure of lifetime reproductive success (lrs) but I don't want to count all the offspring produced over a lifetime by ...
-1
votes
0answers
31 views

Resources for a practical guide to bacterial identification per human specimen

as an instructor in medical microbiology department in a medical college, I need to know what other people use in their practical sessions for post-graduate students as their short yet concise ...
1
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1answer
27 views

Pooling for qPCR

I am comparing miRNA expression levels in 3 different groups but I am low on money and time. I have to get some preliminary results to get the actual research going so I decided to pool my samples and ...
7
votes
1answer
450 views

Extracting cell-free DNA

Has anybody had any success in extracting cell free DNA from plasma without using expensive kits? I've already spent a lot of my own blood trying to use standard phenol/chloroform methods with very ...
2
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0answers
10 views

Stable isotope sample preparation: Bone Collagen

I am performing research in which I need to extract bone collagen from the foot bones of American Beaver (Castor canadensis). I have read through the literature fairly thoroughly on both the ...
2
votes
3answers
409 views

Bacterial cell lysis buffer used in proteomics procedures

What kinds of detergent-free bacterial lysis buffers exist? The proteins we're extracting will be later analyzed by LC-MS/MS, and we're looking for a lysis buffer that won't interfere with this ...
1
vote
1answer
87 views

Help analyzing SDS-Page gel

In this experiment, we transformed a truncation of the NFAT protein sequence into a plasmid vector to be expressed in E.Coli as a fusion protein with GST. We also attempted to transform the normal ...
0
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0answers
26 views

Alternative to Sterilizing Under Pressure

I want to grow mycelium in agar, and all the resources I have found say to sterilize the agar under 15 psi, but I do not have the necessary equipment to sterilize the agar under pressure. Is there an ...
1
vote
2answers
80 views

Getting PCR amplification at annealing higher than Tm!

I am amplifying a gene where in a gradient pcr i am getting amplification at an annealing temperature about 5 degrees (67) higher than Tm (62.5)? What is wrong here? Also, I am getting a very strong ...
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0answers
34 views

My flow cytometry antibody won't reach saturation

I am staining human neutrophil populations (CD177) and then trying to get a different test target antibody to give a dose response to binding to this population. I have tried concentrations of the ...
4
votes
1answer
122 views

Good pipetting technique?

Good pipetting technique is essential for many biologists, but it can be hard to get right. When I take 1 µl of liquid using a micropipette, I seem to always take less than 1 µl, and that amount is ...
3
votes
0answers
17 views

How would one identify cellular transcription factors associated with a viral protein in a treated cell line?

I've been working as the computer guy for a microbiology lab for the past few months. I've always been interested in bench work, but my wet lab experience is rather limited and thus so is my ...
3
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1answer
35 views

Why is it advised to avoid bubble formation during mixing?

I have been told not to vortex solution containing protein. The reason I was given is bubble formation. Here I am interested in the effect of bubble formation in general.
2
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3answers
410 views

Amplification technique for proteins similar to PCR for DNA?

I know PCR can be used to amplify a tiny sample of DNA in order to perform experiments. Is there a similar technique to use on a protein sample? More specifically, I'm not interested in "cutting" up ...
0
votes
1answer
47 views

probe amplification in MLPA

I'm reading an article about MLPA (Multiplex ligation-dependent probe amplification) and I got stuck on this sentence: The advantage of splitting the probe into two parts is that only the ...
1
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0answers
49 views

Making mistakes in the laboratory [closed]

I have just started an internship in a microbiology laboratory. I have had a rather small amount of laboratory experience before this so had to learn mostly everything from scratch. I have been there ...
4
votes
1answer
555 views

For people who work in microbiology labs, what software do you use to manage your strains?

It's just a small microbiology lab that currently records everything on paper, and there's quite few mutants as well. Is Excel commonly used for this sort of thing? Or is there a better software to ...
3
votes
1answer
46 views

Resolution of X-ray crystallography

A structure determined by X-ray crystallography has a resolution of 1.5 Å. When I look at the coordinates, I find every backbone C-N distance is 1.32 Å.i.e. Accurately predicted. If resolution is not ...
12
votes
1answer
5k views

When running gels what is the difference between constant volts or constant amps?

In general, you want to be consistent with running your gels either at constant volts or constant amps. However, it is very clear that during the progression of both PAGE and agarose gels, the free ...
0
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0answers
22 views

What Helminth species has the simplest LABORATORY CONDITION for hatching?

I am very interested to hatch the eggs of a parasite in LABORATORY ! but for my first experience I need a species that is not so expensive or time consuming for experiment and can be done by an ...
3
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0answers
50 views

Why do I get such a strong background in a detection of DIG-labeled DNA (Southern Blot)?

I have done a Southern Blot Analysis of DNAmt transferred to nylon membranes. The DNA was firstly loaded on a 2% agarose gel. An immunoassay was done to detect the bands using Anti-Digoxigenin ...
3
votes
1answer
123 views

In CRISPR bacteria, how does viral genomes get integrated into the spacers of CRISPR? Also, in its use, where does Cas9 cut the DNA?

I've been out of Biology for about a year polishing my programming skills. I know CRISPR/Cas9 allows targeted 'cutting' of DNA via RNA-guidance. Few questions regarding this. Regarding to its ...
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0answers
67 views

Why do PCR manufacturers recommend assembling the reaction mix on ice?

Many PCR manufacturers recommend preparing the reaction mix for the reaction on ice. For example, here is NEB's recommendation for their Q5 polymerase. Other manufacturers also include similar steps: ...
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vote
1answer
122 views

Can you measure plasma glucose with a regular glucose meter?

I'm wondering if plasma glucose can be measured with a regular glucose meter and strips like this. I know these meters are normally used to measure whole blood glucose, but can they measure plasma ...
0
votes
1answer
36 views

what benefits does lab procedure Realtime PCR have in gene silencing experiments? [closed]

RT-PCR performed in gene silencing but mechanistically what are the benefits of this procedure?
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0answers
20 views

Extraction of RNA from algae

I have used a standard protocol (I will give the bibliography below) to extract RNA from an algae (Posidonia) but I have get literally nothing, since I could not even see traces of the two rRNA. I ...
3
votes
1answer
60 views

Is there a difference between Luria Broth and Lysogeny Broth?

Is there a difference between Luria Bertani and Luria Broth? Or are they both the same thing? Is it necessary to autoclave LB medium after it has been made?
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11 views
4
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1answer
190 views

How sterile is working next to a bunsen burner?

When I was still doing lab work, many people would just wear gloves and work next to a bunsen burner because the clean benches were all in use. This was mostly for plating bacteria like Bacillus ...
2
votes
1answer
231 views

How long can a human neuron live outside the body in a controlled environment?

Have there been any experiments that have kept neurons alive (stationary), without preserving methods such as freezing? If yes, then how long were the cells kept alive for?
0
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0answers
12 views

Easiest way to digitally store pathlab reports,blood reports and other reports

I'm engineering student, and working on cheap way to store medical records of a person. I want to know way to partially digitize variety of reports, by partial I mean that I would store photo of ...
3
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0answers
26 views

How do I test if microbes have survived after dehydration?

I have a solution containing various bacteria and fungi. My aim is to place solution on filter paper, and wait until it dries. I then wish to test if the organisms have survived, either on the dried ...
1
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0answers
26 views

Batch several sequences for absent restriction sites

I have a collection of about 120 7kB sequences I would like to check for ether a list of specific restriction sites, or what restriction sites might be absent in all of them. Is there a app or ...
1
vote
1answer
43 views

Making positive charged polyacrylamide

I am interested in positive charged polyacrylamide to electrophorese molecules I am interested in. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2643323/ ...
0
votes
1answer
58 views

On which amino-acids residue is the SDS acting on?

I would like to know exactly what is the mechanisme of the SDS, and I would like to know on which amino-acids residue the SDS is acting on. Can you help me please ? Thank you in advance !
11
votes
3answers
535 views

How can I label cryotubes in a way that eliminates the problem of legible hand-writing?

My lab stores biological material (tissue, cells, plasma, serum) in a -130 C, liquid nitrogen freezer. The cryotubes that we use to store a samples are labelled by hand which frequently creates ...
7
votes
2answers
379 views

What is the best way to express two proteins in a mammalian cell?

I have two proteins and I will be preparing a vector with both genes for stable transfection. Each protein will have their own promoter and I will use piggyBac vector to insert a single cassette with ...
0
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0answers
16 views

Quantitative measurement of taxis

Is there a way to quantitatively measure different forms of taxis, like chemotaxis, electrotaxis, phototaxis, etc. Along with that, how can we also quantitatively compare the strengths of the ...
2
votes
1answer
51 views

Where does the inverse seconds unit come from in the association constant?

I'm working to determine Kd(s) kinetically by generating association and disassociation curves. Kon (association constant, or on-rate) is in inverse seconds multiplied by inverse molar. I get that ...
3
votes
1answer
72 views

How C. Elegans is used for siliencing genes

The experiment that is using C. Elegans to silence the Genes. I have a question about Why and how C. Elegans can use the DNA plasmid that is generated with the gene of interest in the bacteria by ...
5
votes
1answer
130 views

Normal ECG/EKG Measurement?

I'm getting started using ECG using a 3 lead system by iWorx. I have place the leads on my two wrists and the ground on my ankle and have recorded some data into the provided LabScribe 3 software ...
2
votes
3answers
199 views

Is RNase AWAY in the lab dangerous?

I use RNase AWAY in the lab. I would like to know how dangerous this chemical is for health. For example, when I remove my gloves my hands smell because of the RNAse AWAY
3
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0answers
247 views

How much salt [NaCl] is too much in DNA precipitation?

In DNA extractions, how much is too much salt in a CTAB extraction buffer? Protocols hover around 2.5 molar; if you go over this (e.g. 25 molar), will you saturate your solution, and precipitate the ...