Biochemistry and Molecular Biology

Hello, As you've might have noticed most enzymes on the market are ridiculously expensive compared to production costs. What I have heard from people I've worked with is that it semi legal to isolate enzymes (excluding high fidelity versions) if it is used for non commercial experiments. Besides I have never heard of a company going after a single person for producing enzymes, they concentrate on Universities and Institutes. With that said I wanted to ask if you guys have any good protocols for isolating polymerases, ligases, Dna Ladders, restriction endonucleases or any other enzyme for the home laboratory. thanks in advance

Hi, What's the difference between a total cell lysate and a whole cell lysate? glo

Hi, I wish to take a supplement containing lutein, zeaxanthin, and meso zeaxanthin, but there only seems to be three companies that make it. I have been in touch with researchers in Ireland who have been researching the benefits of a brand by the name of MacuShield. The possible benefits indeed do seem to be significant for those afflicted with macular degeneration. My problem is that all three companies' products are encapsulated by gelatine, which is unacceptable for a vegetarian like myself. I have been in touch with the aforementioned researchers, and with another company by the name of MacuHealth, and I am getting conflicting messages regarding whether or no…

Hi all, Please I'm having some very serious problems on my project. What I am trying to do is to: 1. extract an EGFP gene from pSAT1-EGFP-C1 (4553 bp) and clone it onto the binary vector, pPZP-RCS2-ocs-bar-R1 (8637 bp)and then transform it with competent cells. 2. Later on, I will use it to clone some genes from tomato plants by silencing usin VIGS. But the problem I am having now is in the first step. I havn't been able to clone the EGFP gene onto the binary vector. The protocol I have been using is: For the vector (a) plasmid extraction of vector (plasmid conc. is usually high) (b) enzyme digestion of vector with Asc1 enzyme (I get one band after cutting)…

hi all! i want to use chelex 100 sodium form (Sigma 50-100 dry mesh) in order to remove calcium traces from my protein but i dont know how to use it. any suggestion is always wellcome. cheers, s

hi, Can any one help me in deciphering the mistake in processing adult mouse skin and intestine for In Situ Hybridisation and histological staining. I am Fixing my tissue in 4% PFA overnight at 4 degree and then dehydrating in graded series of alcohol (25%,50%, 70%,80%,90% and 100%). Xylene is used as clearing for 3 minutes and 2min and then paraffin washes of 1 hour and overnight duration at 65degree, following embedding in paraffin. I am not getting good quality sections (5-7micro meters), tissue gets torn off from between the sections. Skin tissue section fall off from the slide during In Situ Hybridisation process.

Hello, I am currently reconstituting the light activated proton pump bacteriorhodopsin into giant unilamellar vesicles. It is important that I dissolve the protein in diethyl ether in order for the procedure to work. I was wondering if it is safe to make a stock solution to store bacteriorhodopsin in for an extended period of time(on the order of months)? If anyone has any insight on what factors might come into play or if they know from experience that it is safe or not it would be greatly appreciated. Thanks, Adam

Hi, How long the protein samples( cell Lysate) can be stored in 4 degree? If its there at 4 degree for more than 12 hours won't the Proteins be degraded?Can it be still used for the experiments like WB? Alamelu

Hi everyone In my research I produce plasmidial DNA-PEI complexes, which later I entrap into anionic liposomes. In order to quantify the entrapped DNA, I ultra-centrifuge the liposomes, then re-suspend them in 0.1% Triton X100 for 30 minutes to break down the cells and then add Heparin to 0.3% for another 30 minutes in order to release the DNA from the PEI. I then quantify the DNA using PicoGreen. In addition, I quantify the DNA that was not entrapped, in the supernatant from the ultracentrifuge. The problem is, the DNA in the liposomes plus the DNA in the supernatant don't add up to the amount of the DNA I added in the first place! Is there a problem in my met…

Greetings all, I plan to perform Western Blot on Human Hair Follicle Dermal Papilla Cells. But this is my first time performing Western Blot, I hardly know what to choose / prepare for the procedure. Specifically, I wanted to detect these proteins: ERK 1/2 Phospho-ERK 1/2 Akt Phospho-Akt Bcl2 Beta-Catenin FGF-7 As for the procedure, I am going to use RadioImmunoPrecipitation Assay buffer to lyse the cells. For the Antibodies, I've gotten a list of Antibodies for each of the proteins: ERK 1/2: www.rndsystems.com/pdf/mab1576.pdf - Mouse Anti-Human ERK 1/2, 100 ug Phospho-ERK 1/2: http://www.rndsystem...pdf/MAB1825.pdf - Mouse Anti-Human phospho-ERK 1/…

It's supposed to be a trisaccharide but couldn't find anything about it from google.

Hi, need a little help here. If you are assaying the pressence of a protein during several stages of development of an animal, and you see that, let's say, P1 and P8 have two bands (let's say P0 is the embryonic stage and no protein iband is seen; and the latter stages are P1 to P9) and the rest (P2 to P7 and P9) one band. What is the most probable explanation to this? And how can you verify it? This is what I think: 1)the antibody is recognizing more than one epitope (not very specific) or 2) there is a protein with a very similar epitope (not really think is probable, also relates to 1) in some stages or 3) the protein is an oligomer and it is cleaved in some …

I need to come up with a reason why an extra gene that codes for an MFS transporter in bacteria would increase colonization in the rizosphere of a plant. So far all I've come up with "the cells can respire better, so it colonizes better". How would I start making more connections? This isn't a test question, it's just a for a paper so any old reason that I can back up and say "disprove this" will suffice. edit: I should note, the gene seems to be unique to this individual, and the individual was selected because of it's rizosphere colonization properties.

Hi. Passing an electric current, even for a brief second, has devastating effects on living cells, destroying/killing them. If a person has a topical infection, would applying electrical current confined to a short path within the affected area with reduced/minimal effect to healthy surrounding tissues work ? Does topical anesthesia work for minor electroshocks ? Does healthy skin, muscle cells killed in an electroshock get replaced by new growth ? Electrodes placement would need some thoughtful technique, but nothing seems extraordinary there. Thanks for opinions. -Please move post to proper section of the forum if this is not the correct one-

Hello, I was wondering if someone could help me. After weeks of research it is driving me crazy! Any help would be greatly, greatly, greatly appreciated! I am looking for someone that could help me understand the chemical reactions and processes of fruit and vegetable preservation and could suggest processes that will work with what I am trying to create. I am a artist that is working with fruits and vegetables who is trying to create vegetable and fruit paper bowls here are some examples: link to images I have done some research on line and I am still having problems understanding the preservation of the organic materials and color. I have looked into food…

i have a molecular biology presentation due next week so i want something new to talk about to impress my doctor ideas please ?

Now I'm at high school. And yesterday, I've learned that respiration is c6 o6 h12 broken by o2 to give co2 + h20 + energy. If the main content of glucose is carbohydrate(coh) then can we react water and c02 to get coh in any form? If we could get that, then we could just make an artificial canal from lungs to stomach. Please think about it and give me some explanative reply because i am only just trying to enter the world of biology.

First post, hello to everybody! I am interested in the mechanisms determining fat distribution in the body. Is there a documented difference in white adipose tissue in various location of the body? There are countless studies on the association of fat distribution and other factors, but very little information about molecular mechanisms and causality. Any help / reading suggestions are welcome.

Hi, I´ve been having issues with polystyrene Functional Beads for Cytometric Bead Array from BD Bioscience. In my experiment, I covalently link water-soluble proteins to the surface of these functional beads and I have some troubles with storage. When I do cytometric analysis it seems like the beads fell apart. Does anyone here have any idea what to use to preserve them? Thanks in advance .

I'm a college sophomore and I'm looking into DNA methylation, especially how it is maintained through cell generations. I've been doing some research online and from that it seemed to me that scientists had a pretty good idea of how to accurately detect methylation patterns. However, I came across an article titled " Cooperativity between DNA Methyltransferases in the Maintenance Methylation of Repetitive Elements" which claimed that until their experimental design, no method to analyze the levels of hemimethylation at individual regions in the genome existed. This did not seem to match up with the rest of my research; how can the accuracy of maintenance methylation …

Hi People, Is it possible to label the N terminal of a peptide with Rhodamine B. As u know Rhodamine B has a carboxylic acid and under coupling conditions, that is HBTU/HOBt and DIPEA in DMF, could be easily incorporated. This is what I guess...but I wanted to ask whether u have any experience in the incorporation of this fluorophore to any peptide sequence. I didn't find any protocol in which is reported...and therefore I feel a little bit afraid about if it is possible to attach it... Thanks, Verbatim:25

Vladimir Matveev. Protoreaction of Protoplasm. Cell. Mol. Biol. 51(8): 715-723, 2005 Abstract. My goal is to describe briefly the universal cellular reaction (UCR) to external actions and agents. This general reaction was the main subject of investigation by the scientific school of the outstanding Russian cytologist, Dmitrii Nasonov (1895-1957). The UCR consists of two phases of complex changes in cellular viscosity and turbidity, in the cell's ability to bind vital dyes, in the resting membrane potential, and in cellular resistance to harmful actions. Works from the Nasonov School have shown that these changes are based on structural-functional transformations of ma…

This may sound like a strange question, but due to my lack of cellular biology I can't seem to determine the feasibility of such a question: Is it possible to inject chloroplasts into an animal cell and force symbiosis, and thus allowing it photosynthesize and respirate without forming cell walls and the big ass vacuoles that plants have? I also realized that the replication of the organelles require additional genetic information to be injected into various chromosomes, so any thoughts?

guys, I'd like to discuss this idea (on youtube on the link below) to record which molecule(s) is(are) interacting with which ones when two kind of cells met. So far, in any case, at least one of the partner has to be know. The game then consist to try to identify who is binding to it (blotting, screening, labeling). Most of the time, it required firstly purification of known proteins. Here the idea is to reverse the mapping of cell-cell, cell-protein and protein-protein interaction by freezing, linking them with UV and then sequence the attached parts after isolation to identify the interacting partners. I use freeze clamp to freeze ionic channel so, they can be…

Hi, I have searched, but only found it's applications, so I was wondering if someone can please explain me what that is.